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PubMed2006

Preparation of poly (alkylcyanoacrylate) nanoparticles by polymerization of water-free microemulsions.

Phase diagrams of the pseudoternary systems ethyloleate, polyoxyethylene 20 sorbitan mono-oleate/sorbitan monolaurate and propylene glycol with and without butanol as a co-surfactant were prepared. Areas containing optically isotropic, one-phase systems were identified and samples therein designated as droplet, bicontinuous or solution type microemulsions using conductivity, viscosity and self-diffusion NMR. Nanoparticles were prepared by polymerization of selected microemulsions with ethyl-2-cyanoacrylate and the morphology of the particles was investigated. Addition of monomer to all types of microemulsions led to the formation of nanoparticles, which had an average size of 244 +/- 25 nm, an average polydispersity index of 0.15 +/- 0.04 and a zeta-potential of -17 +/- 3 mV. The formation of particles from water-free microemulsions of different types is surprising, particularly considering that polymerization is expected to occur at a water-oil interface by base-catalysed polymerization. It would appear that propylene glycol is sufficiently nucleophilic to initiate the polymerization. The use of water-free microemulsions as templates for the preparation of poly (alkylcyanoacrylate) nanoparticles opens up interesting opportunities for the encapsulation of bioactives which do not have suitable properties for encapsulation on the basis of water-containing microemulsions.

Cyanoacrylates↗
PubMed1999

Binding of polymerizing fibrin to integrin alpha(IIb)beta(3) on chymotrypsin-treated rabbit platelets decreases phosphatidylinositol 4,5-bisphosphate and increases cytoskeletal actin.

Incubation of polymerizing fibrin with washed, chymotrypsin-treated (CT) rabbit platelets resulted in the formation of platelet-fibrin clots, a decrease of 27.4% (P < 0.001, n = 20) in the amount of phosphatidylinositol 4,5-bisphosphate (PIP) and an increase of 37.2% (P < 0.001, n = 20) in phosphatidylinositol 4-phosphate (PIP(2)), apparently because of a shift in the equilibria between PIP(2) and PIP toward PIP. In contrast, incubation of fibrinogen with CT-platelets resulted in agglutination of platelets, but no changes in the phosphoinositides. Preincubation of CT-platelets with staurosporine (1 microM) to inhibit protein phosphorylation, okadaic acid (1 microM) to inhibit protein phosphatases, genistein (100 microM) to inhibit protein tyrosine phosphorylation, PGE (10 microM) to increase cAMP and cause Ca2+ sequestration or with wortmannin (50 nM) to inhibit phosphoinositide 3-kinase, did not inhibit the polymerizing fibrin-induced decrease in PIP2 and increase in PIP. Preincubation with cytochalasin E (CE, 5 microM) inhibited the decrease in PIP(2) by 57% (P < 0.01, n = 8), but not completely. CE did not affect the resting levels of PIP2. Thus, the state of the actin cytoskeleton appears to affect signalling from the integrin receptor alpha(IIb)beta(3) to the enzymes of phosphoinositide interconversion. Platelet cytoskeleton actin content increased by 16.4 4.1% (P < 0.01, n = 5) because of polymerizing fibrin binding to CT-platelets. This is the first demonstration of a pathway involving a decrease in PIP(2) caused by binding of polymerizing fibrin to alpha(IIb)beta(3) and an associated increase in cytoskeletal actin, which may be involved in reorganization of the cytoskeleton for platelet-mediated clot retraction.

Journal Article↗
PubMed2001

Synthesis and evaluation of a nonsteroidal anti-inflammatory polymeric prodrug for sustained and site-specific delivery.

A new polymerizable drug derivative of diclofenac sodium was synthesized and characterized in terms of melting point, elemental analysis, and infrared spectroscopy. It was then polymerized to obtain a new polymeric prodrug. The prodrug was evaluated for its viscosity, drug content, and in vitro drug release behavior at pH 1.2 and 7.2. The in vitro studies showed that the drug release takes place predominantly at the higher pH and in a sustained manner, as hypothesized. Stability at room temperature, bioavailability, and ulcer-inducing effect of the polymeric prodrug were also studied. The investigations showed complete drug absorption from the polymeric prodrug with a statistically significant decrease in ulcer scoring effect, thus showing its potentialfor site-specific and sustained drug delivery.

Animals↗
PubMed2003

Organic redox-initiated polymerization process for the fabrication of hydrogels for colon-specific drug delivery.

Organic-redox initiated polymerization technique based on the co-initiators system comprising benzoyl peroxide and N-phenyldiethanolamine was used at ambient temperature to fabricate pH-responsive hydrogels. The effects of changes in the concentration of the co-initiators system, the ratio in which the co-initiators combined, the type of the polymerization solvent, the pH of the hydrating medium, the concentration of the cross-linking agent based on azo-bond and the pH-sensitive cross-linking agent on the properties of the hydrogels were investigated. Increasing the concentration of the co-initiators system, decreasing the concentration of the two types of cross-linking agents, and replacing DMSO by ethanol as the polymerization solvent resulted in hydrogels with increased equilibrium swelling ratio and increased molecular weight between cross-links at pH 7.4. Increasing the concentration of N-phenyldiethanolamine while keeping the concentration of benzoyl peroxide constant gave hydrogels with increased equilibrium swelling ratios. The equilibrium swelling ratios of the hydrogels at pH 2.0 were not affected by the factors investigated. The polymerization technique may be suitable for the design of drug delivery systems containing thermolabile bioactive agents like peptides and proteins.

Azo Compounds↗
PubMed2003

Preparation and characterization of diacetylene polymerized liposomes for detection of autoantibodies.

Affinity diacetylene liposomes were prepared with 10,12-tricosadiynoic acid and cardiolipin as the affinity ligand for anticardiolipin antibodies at a molar ratio of 80:20. Polymerization was carried out under UV irradiation, and the color transitions were monitored by visible absorption spectroscopy. Peaks at 635 nm (blue form), 540 nm (purple form), and 480 nm (red form) were observed as a function of time. These polymerized liposomes were used in a noncompetitive immunoassay for detection of anticardiolipin antibodies. Color changes were observed when reference serum containing specific immunoglobulin G, IgG, was added to polymerized liposome dispersions. The colorimetric signal due to IgG adsorption on the liposome surface was quantified as a colorimetric response defined as the change in percentage of blue color related to the initial percentage of blue color in the solutions. The colorimetric response was 10 times higher for specific IgG compared with nonspecific ones. These results suggest the unique potentialities of affinity diacetylene polymerized liposomes in the development of biosensors for diagnosis of autoimmune diseases.

Acetylene↗
PubMed1985

Relationship of actin polymerization and depolymerization to light scattering in human neutrophils: dependence on receptor occupancy and intracellular Ca++.

When exposed to the N-formylated chemoattractant peptides, neutrophils undergo a transient ruffling followed by a polarization that involves a redistribution of F-actin (Fechheimer, M., and S. H. Zigmond, 1983, Cell Motil., 3:349-361). The cells also undergo a biphasic right angle light scatter response whose first phase is maximal 10-15 s after exposure to the stimulus, and whose second phase is longer in duration and maximal only after 1 min or more (Yuli, I., and R. Snyderman, 1984, J. Clin. Invest. 73:1408-1417). We now report that the first phase is accompanied by a transient polymerization of actin (monitored by cytometric analysis of phallacidin staining according to the method of Howard, T. H., and W. H. Meyer, 1984, J. Cell Biol., 98:1265-1271) and the second phase is accompanied by a more sustained polymerization of actin. Based on correlated measurements of ligand binding (Sklar, L. A., D. A. Finney, Z. G. Oades, A. J. Jesaitis, R. G. Painter, and C. G. Cochrane, 1984, J. Biol. Chem., 259:5661-5669) and intracellular Ca++ elevation (under conditions where we use the fluorescent Ca++ chelator Quin 2 to modulate intracellular Ca++ levels), we conclude that this first phase requires less than 100 receptors/cell (out of 50,000) and does not require the release of intracellular stores of Ca++. In contrast, the sustained polymerization requires both the occupancy of thousands of receptors (an estimated 10% of the receptors per minute) and may be somewhat sensitive to the availability of intracellular Ca++. When ligand binding is interrupted, F-actin rapidly depolymerizes with a half-time of no greater than approximately 15 s, and the transient light scatter response decays toward its initial value in parallel. Partial disaggregation of the cells follows the recovery of these responses. Based on these observations, we suggest that transient actin polymerization and transient cell ruffling give rise to transient aggregation as long as degranulation is limited.

Actins↗
PubMed1988

Polymerization of actin by positively charged liposomes.

By cosedimentation, spectrofluorimetry, and electron microscopy, we have established that actin is induced to polymerize at low salt concentrations by positively charged liposomes. This polymerization occurs only at the surface of the liposomes, and thus monomers not in direct contact with the liposome remain monomeric. The integrity of the liposome membrane is necessary to maintain actin in its polymerized state since disruption of the liposome depolymerizes actin. Actin polymerized at the surface of the liposome is organized into two filamentous structures: sheets of parallel filaments in register and a netlike organization. Spectrofluorimetric analysis with the probe N-pyrenyl-iodoacetamide shows that actin is in the F conformation, at least in the environment of the probe. However, actin assembly induced by the liposome is not accompanied by full ATP hydrolysis as observed in vitro upon addition of salts.

Actins↗
PubMed1989

Guanine nucleotide-induced polymerization of actin in electropermeabilized human neutrophils.

The effects of exogenous guanine nucleotides on the polymerization of actin in human neutrophils were tested in an electropermeabilized cell preparation. Close to 40% permeabilization was achieved with a single electric discharge as measured by nucleic acid staining with ethidium bromide or propidium iodide with minimal (less than 2%) release of the cytoplasmic marker lactate dehydrogenase. In addition, electropermeabilized neutrophils retained their capacity to produce superoxide anions and to sustain a polymerization of actin in response to surface-receptor dependent stimuli such as chemotactic factors. Electropermeabilization produced a rapid and transient permeabilization that allowed the entry of guanine nucleotides into the cells. GTP and, to a larger extent, its nonhydrolyzable analog guanosine 5'-O-2-thiotriphosphate (GTP[S]), induced a time- and concentration-dependent polymerization of actin, as determined by increased staining with 7-nitrobenz-2-oxa-1,3-diazolylphallacidin. The effects of the aforementioned guanine nucleotides were antagonized by GDP[S], but were insensitive to pertussis toxin. Cholera toxin potentiated to a small degree the amount of actin polymerization induced by GTP[S]. These results provided direct evidence for the involvement of GTP-binding proteins in the regulation of the organization of the cytoskeleton of neutrophils, an event that is of crucial importance to the performance of the defense-oriented functions of these cells.

Actins↗
PubMed1990

Exogenous nucleation sites fail to induce detectable polymerization of actin in living cells.

Most nonmuscle cells are known to maintain a relatively high concentration of unpolymerized actin. To determine how the polymerization of actin is regulated, exogenous nucleation sites, prepared by sonicating fluorescein phalloidin-labeled actin filaments, were microinjected into living Swiss 3T3 and NRK cells. The nucleation sites remained as a cluster for over an hour after microinjection, and caused no detectable change in the phase morphology of the cell. As determined by immunofluorescence specific for endogenous actin and by staining cells with rhodamine phalloidin, the microinjection induced neither an extensive polymerization of endogenous actin off the nucleation sites, nor changes in the distribution of actin filaments. In addition, the extent of actin polymerization, as estimated by integrating the fluorescence intensities of bound rhodamine phalloidin, did not appear to be affected. To determine whether the nucleation sites remained active after microinjection, cells were first injected with nucleation sites and, following a 20-min incubation, microinjected with monomeric rhodamine-labeled actin. The rhodamine-labeled actin became extensively associated with the nucleation sites, suggesting that at least some of the nucleation activity was maintained, and that the endogenous actin behaved in a different manner from the exogenous actin subunits. Similarly, when cells containing nucleation sites were extracted and incubated with rhodamine-labeled actin, the rhodamine-labeled actin became associated with the nucleation sites in a cytochalasin-sensitive manner. These observations suggest that capping and inhibition of nucleation cannot account for the regulation of actin polymerization in living cells. However, the sequestration of monomers probably plays a crucial role.

Actins↗
PubMed1991

Control of actin polymerization in live and permeabilized fibroblasts.

We have investigated the spatial control of actin polymerization in fibroblasts using rhodamine-labeled muscle actin in; (a) microinjection experiments to follow actin dynamics in intact cells, and (b) incubation with permeabilized cells to study incorporation sites. Rhodamine-actin was microinjected into NIH-3T3 cells which were then fixed and stained with fluorescein-phalloidin to visualize total actin filaments. The incorporation of newly polymerized actin was assayed using rhodamine/fluorescein ratio-imaging. The results indicated initial incorporation of the injected actin near the tip and subsequent transport towards the base of lamellipodia at rates greater than 4.5 microns/min. Furthermore, both fluorescein- and rhodamine-intensity profiles across lamellipodia revealed a decreasing density of actin filaments from tip to base. From this observation and the presence of centripetal flux of polymerized actin we infer that the actin cytoskeleton partially disassembles before it reaches the base of the lamellipodium. In permeabilized cells we found that, in agreement with the injection studies, rhodamine-actin incorporated predominantly in a narrow strip of less than 1-microns wide, located at the tip of lamellipodia. The critical concentration for the rhodamine-actin incorporation (0.15 microM) and its inhibition by CapZ, a barbed-end capping protein, indicated that the nucleation sites for actin polymerization most likely consist of free barbed ends of actin filaments. Because any potential monomer-sequestering system is bypassed by addition of exogenous rhodamine-actin to the permeabilized cells, these observations indicate that the localization of actin incorporation in intact cells is determined, at least in part, by the presence of specific elongation and/or nucleation sites at the tips of lamellipodia and not solely by localized desequestration of subunits. We propose that the availability of the incorporation sites at the tips of lamellipodia is because of capping activities which preferentially inhibit barbed-end incorporation elsewhere in the cell, but leave barbed ends at the tips of lamellipodia free to add subunits.

Actins↗
PubMed1993

Yeast actin with a mutation in the "hydrophobic plug" between subdomains 3 and 4 (L266D) displays a cold-sensitive polymerization defect.

Holmes et al. (Holmes, K. C., D. Popp, W. Gebhard, and W. Kabsch. 1990. Nature [Lond.] 347: 44-49) hypothesized that between subdomains 3 and 4 of actin is a loop of 10 amino acids including a four residue hydrophobic plug that inserts into a hydrophobic pocket formed by two adjacent monomers on the opposing strand thereby stabilizing the F-actin helix. To test this hypothesis we created a mutant yeast actin (L266D) by substituting Asp for Leu266 in the plug to disrupt this postulated hydrophobic interaction. Haploid cells expressing only this mutant actin were viable with no obvious altered phenotype at temperatures above 20 degrees C but were moderately cold-sensitive for growth compared with wild-type cells. The critical concentration for polymerization increased 10-fold at 4 degrees C compared with wild-type actin. The length of the nucleation phase of polymerization increased as the temperature decreased. At 4 degrees C nucleation was barely detectable. Addition of phalloidin-stabilized F-actin nuclei and phalloidin restored L266D actin's ability to polymerize at 4 degrees C. This mutation also affects the overall rate of elongation during polymerization. Small effects of the mutation were observed on the exchange rate of ATP from G-actin, the G-actin intrinsic ATPase activity, and the activation of myosin S1 ATPase activity. Circular dichroism measurements showed a 15 degrees C decrease in melting temperature for the mutant actin from 57 degrees C to 42 degrees C. Our results are consistent with the model of Holmes et al. (Holmes, K. C., D. Popp, W. Gebhard, and W. Kabsch. 1990. Nature [Lond.]. 347:44-49) involving the role of the hydrophobic plug in actin filament stabilization.

Actins↗
PubMed1999

Laminin polymerization induces a receptor-cytoskeleton network.

The transition of laminin from a monomeric to a polymerized state is thought to be a crucial step in the development of basement membranes and in the case of skeletal muscle, mutations in laminin can result in severe muscular dystrophies with basement membrane defects. We have evaluated laminin polymer and receptor interactions to determine the requirements for laminin assembly on a cell surface and investigated what cellular responses might be mediated by this transition. We found that on muscle cell surfaces, laminins preferentially polymerize while bound to receptors that included dystroglycan and alpha7beta1 integrin. These receptor interactions are mediated through laminin COOH-terminal domains that are spatially and functionally distinct from NH2-terminal polymer binding sites. This receptor-facilitated self-assembly drives rearrangement of laminin into a cell-associated polygonal network, a process that also requires actin reorganization and tyrosine phosphorylation. As a result, dystroglycan and integrin redistribute into a reciprocal network as do cortical cytoskeleton components vinculin and dystrophin. Cytoskeletal and receptor reorganization is dependent on laminin polymerization and fails in response to receptor occupancy alone (nonpolymerizing laminin). Preferential polymerization of laminin on cell surfaces, and the resulting induction of cortical architecture, is a cooperative process requiring laminin- receptor ligation, receptor-facilitated self-assembly, actin reorganization, and signaling events.

Actins↗
PubMed1978

Polymerization of actin. IV. Role of Ca++ and H+ in the assembly of actin and in membrane fusion in the acrosomal reaction of echinoderm sperm.

When Pisaster, Asterias, or Thyone sperm are treated with the ionophore A23187 or X537A, an acrosomal reaction similar but not identical to a normal acrosomal reaction is induced in all the sperm. Based upon the response of the sperm, the acrosomal reaction consists of a series of temporally related steps. These include the fusion of the acrosomal vacuole with the cell surface, the polymerization of the actin, the alignment of the actin filaments, an increase in volume, an increase in the limiting membrane, and changes in the shape of the nucleus. In this report, we have concentrated on the first two steps in this sequence. Although fusion of the acrosomal vacuole with the cell surface requires Ca++, we found that the polymerization of actin instead appears to be dependent upon an increase in intracellular pH. This conclusion was reached by applying to sperm A23187, X537A, or nigericin, ionophores which all carry H+ at high affinity, yet vary in their affinity for other cations. When sperm are suspended in isotonic NaCl, isotonic KCl, calcium-free seawater, or seawater, all at pH 8.0, and the ionophore is added, the actin polymerizes explosively and an efflux of H+ from the cell occurs. However, if the pH, of the external medium is maintained at 6.5, the presumed intracellular pH, no effect is observed. And, finally, if egg jelly is added to sperm (the natural stimulus for the acrosomal reaction) at pH 8.0, H+ is also released. On the basis of these observations and those presented in earlier papers in this series, we conclude that a rise in intracellular pH induces the actin to disassociate from its binding proteins. Now it can polymerize.

Acrosome↗
PubMed1979

Polymerization of actin. VI. The polarity of the actin filaments in the acrosomal process and how it might be determined.

The polarity of the actin filaments which assemble from the nucleating body or actomere of Thyone and Pisaster sperm was determined using myosin subfragment 1 decoration. The polarity was found to be unidirectional with the arrowheads pointing towards the cell center. When polymerization is induced at low temperature with concentrations of actin near the critical concentration for polymerization, elongation of filaments occurs preferentially off the apical end. If the sperm are induced to undergo the acrosomal reaction with an ionophore, the polarity of the actin filaments attached to the actomere is the same as that already described, but the filaments which polymerize parallel to, but peripheral to, those extending from the actomere are randomly polarized. These randomly polarized filaments appear to result from spontaneous nucleation. When sperm are induced to undergo the acrosomal reaction with eggs, the polarity of the actin filaments is also unidirectional with the arrowheads pointing towards the cell center. From these results we conclude: (a) that the actomere, by nucleating the polymerization of actin filaments, controls the polarity of the actin filaments in the acrosomal process, (b) that the actomere recognizes a surface of the actin monomer that is different from that surface recognized by the dense material attached to membranes, and (c) that egg myosin could not act to pull the sperm into the egg. Included is a discussion of how the observation that monomers add largely to one end of a decorated filament in vitro relates to these in vivo observations.

Acrosome↗
PubMed1980

Cytochalasins inhibit nuclei-induced actin polymerization by blocking filament elongation.

Polylysine was found to induce polymerization of muscle actin in a low ionic strength buffer containing 0.4 mM MgCl2. The rate of induced polymerization was dependent on the amount and on the molecular size of the polylysine added. A similar effect was obtained by adding actin nuclei (containing about 2-4 actin subunits) cross-linked by p-N,N'-phenylenebismaleimide to G-actin under the same conditions, suggesting that the effect of polylysine is due to promotion of the formation of actin nuclei. Polymerization induced by polylysine and by cross-linked actin nuclei was inhibited by low concentrations (10(-8)-10(-6)M) of cytochalasins. Binding experiments showed that actin filaments, but not actin monomers, contained high-affinity binding sites for [3H]cytochalasin B (one site per 600 actin monomers). The relative affinity of several cytochalasins for these sites (determined by competitive displacement of [3H]dihydrocytochalasin B) was: cytochalasin D greater than cytochalasin E approximately equal to dihydrocytochalasin B. The results of this study suggest that cytochalasins inhibit nuclei-induced actin polymerization by binding to highly specific sites at the point of monomer addition, i.e., the elongation site, in actin nuclei and filaments.

Actins↗
PubMed1982

Nucleated polymerization of actin from the membrane-associated ends of microvillar filaments in the intestinal brush border.

We examined the nucleated polymerization of actin from the two ends of filaments that comprise the microvillus (MV) core in intestinal epithelial cells by electron microscopy. Three different in vitro preparations were used to nucleate the polymerization of muscle G-actin: (a) MV core fragments containing "barbed" and "pointed" filament ends exposed by shear during isolation, (b) isolated, membrane-intact brush borders, and (c) brush borders demembranated with Triton-X 100. It has been demonstrated that MV core fragments nucleate filament growth from both ends with a strong bias for one end. Here we identify the barbed end of the core fragment as the fast growing end by decoration with myosin subfragment one. Both cytochalasin B (CB) and Acanthamoeba capping protein block filament growth from the barbed but not the pointed end of MV core fragments. To examine actin assembly from the naturally occurring, membrane-associated ends of MV core filaments, isolated membrane-intact brush borders were used to nucleate the polymerization of G-actin. Addition of salt (75 mM KCl, 1 mM MgSO4) to brush borders preincubated briefly at low ionic strength with G-actin induced the formation of 0.2-0.4 micron "growth zones" at the tips of microvilli. The dense plaque at the tip of the MV core remains associated with the membrane and the presumed growing ends of the filaments. We also observed filament growth from the pointed ends of core filaments in the terminal web. We did not observe filament growth at the membrane-associated ends of core filaments when the latter were in the presence of 2 microM CB or if the low ionic strength incubation step was omitted. Addition of G-actin to demembranated brush borders, which retain the dense plaque on their MV tips, resulted in filament growth from both ends of the MV core. Again, 2 microM CB blocked filament growth from only the barbed (tip) end of the core. The dense plaque remained associated with the tip-end of the core in the presence of CB but usually was dislodged in control preparations where nucleated polymerization from the tip-end of the core occurred. Our results support the notion that microvillar assembly and changes in microvillar length could occur by actin monomer addition/loss at the barbed, membrane-associated ends of MV core filaments.

Actins↗
PubMed1984

Mechanism of regulation of actin polymerization by Physarum profilin.

Physarum profilin reduces the rates of nucleation and elongation of F-actin and also reduces the extent of polymerization of actin at the steady state in a concentration-dependent fashion. The apparent critical concentration for polymerization of actin is increased by the addition of profilin. These results can be explained by the idea that Physarum profilin forms a 1:1 complex with G-actin and decreases the concentration of actin available for polymerization. The dissociation constant for binding of profilin to G-actin is estimated from the kinetics of polymerization of G-actin and elongation of F-actin nuclei and from the increase of apparent critical concentration in the presence of profilin. The dissociation constants for binding of Physarum profilin to Physarum and muscle actins under physiological ionic conditions are in the ranges of 1.4-3.7 microM and 11.3-28.5 microM, respectively. When profilin is added to an F-actin solution, profilin binds to G-actin which co-exists with F-actin, and then G-actin is dissociated from F-actin to compensate for the decrease of the concentration of free G-actin and to keep it constant at the critical concentration. At the steady state, free G-actin of the critical concentration is in equilibrium not only with F-actin but also with profilin-G-actin complex. The stoichiometry of 1:1 for the formation of complex between profilin and G-actin is directly shown by means of chemical cross-linking.

Actins↗
PubMed1980

Production of predominantly polymeric IgA by human peripheral blood lymphocytes stimulated in vitro with mitogens.

Human peripheral blood lymphocytes (PBL) were cultured for various time periods (up to 8 d) in the presence of pokeweed mitogen (PWM), lipopolysaccharide, or Epstein-Barr virus. Cell-free supernates were fractionated on a standardized ultrogel AcA 22 column and the proportion of polymeric and monomeric IgA was determined by radioimmunoassay. The results demonstrate that PBL stimulated with these mitogens produce IgM and IgG with molecular characteristics identical to those found in serum, but that the IgA produced is predominantly of the polymeric type. To prove that this IgA represented disulfide bond-linked polymers rather than aggregated monomers, we have demonstrated that the high molecular weight IgA (a) maintains its polymeric form upon treatment with acidic buffers, (b) contains J chain, a glycoprotein associated only with polymeric immunoglobulins, and (c) dissociates to the monomeric form upon reduction of disulfide bonds. After 1 wk in culture, approximately 60% of the PWM-stimulated cells that contained IgA were positive for IgA2, whereas 40% were IgA1 positive as determined by immunofluorescence. Therefore, peripheral blood contains a population of lymphocytes with the potential to display, after appropriate stimulation and differentiation, characteristics similar to IgA cells found in external secretory tissues. The demonstration of the presence of such cells in the peripheral circulation suggests that these cells are precursors of IgA-producing plasma cells with the potential to populate mucosal tissues.

Humans↗