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Biomedical subjects

D Malamud

Publications and source records attributed to D Malamud.

At least 19 recordsLinked to original sources
PubMed1997

Oral diagnostic testing for detecting human immunodeficiency virus-1 antibodies: a technology whose time has come.

An oral fluid-based test for antibodies to human immunodeficiency virus (HIV), equivalent to serum in its accuracy but safer and easier to use, is now available in the United States. The development of the oral test involved overcoming technical obstacles to the use of oral fluid as a testing medium, including low immunoglobulin G (IgG) titers, suboptimal assay performance, protease degradation of IgG, high viscosity, and lack of a standardized method of specimen collection, all of which contribute to suboptimal assay performance. The currently available oral HIV test utilizes a collection device to isolate a mucosal transudate component of oral fluid rich in IgG. A vial containing a preservative solution facilitates the transport of stable, low-viscosity specimens to the laboratory for testing with an ELISA and confirmatory Western blot assay, specifically designed for use with oral fluid. Non-HIV medical conditions and oral pathologies do not appear to affect oral test results. Hopefully, the availability of a more patient-friendly, portable diagnostic test for antibodies to HIV will facilitate identification of greater numbers of infected individuals with the ultimate goals of early identification, early treatment, and prevention of disease transmission.

Antibodies, Viral↗
PubMed1997

Human submandibular saliva specifically inhibits HIV type 1.

Studies from a number of laboratories have shown the presence of factor(s) in whole, parotid, and submandibular human saliva capable of inhibiting HIV-1 infectivity in vitro. Data from our laboratory suggested that the level of anti-HIV-1 activity is higher in submandibular than parotid or whole saliva. Previous results obtained with pooled submandibular saliva from seronegative individuals included a filtration step following saliva-virus interaction. In this article, we present data on the HIV-1 inhibitory activity of individual submandibular saliva samples collected from 15 donors. We show that although anti-HIV activity is quantitatively similar in most individuals (9 of 15), some (4 of 15) are much less active than others and some (2 of 15) lack inhibitory activity. We also show that for most individuals the level of anti-HIV inhibitor is similar with or without a filtration step. However, 2 of the 15 samples demonstrated activity only after filtration. The quantitative and qualitative anti-HIV activity of individual saliva samples appeared to reflect differences in the individual donors. We further show that the anti-HIV activity of submandibular saliva is demonstrated not only against laboratory strains of HIV-1 but is similarly active against three clinical HIV-1 isolates. In contrast, submandibular saliva had little effect on the infectivity of HIV-2 or SIV.

Antiviral Agents↗
PubMed1997

The microbicidal agent C31G inhibits Chlamydia trachomatis infectivity in vitro.

Safe and effective vaginal microbicidal compounds are being sought to offer women an independent method for protection against transmission of sexually acquired pathogens. The purpose of this study was to examine the efficacy of two formulations of one such compound, C31G, against Chlamydia trachomatis serovar E alone, its host epithelial cell (HEC-1B) alone, and against chlamydiae-infected HEC-1B cells. Preexposure of isolated, purified infectious chlamydial elementary bodies (EB) to C31G, at pHs 7.2 and 5.7, for 1 h at 4 degrees C resulted in reduced infectivity of EB for HEC-1B cells. Examination of the C31G-exposed 35S-EB on sodium dodecyl sulfate-polyacrylamide gel electrophoresis autoradiographs and by Western blotting revealed a C31G concentration-dependent and pH-dependent destabilization of the chlamydial envelope, resulting in the release of chlamydial lipopolysaccharide and proteins. Interestingly, when the host human genital columnar epithelial cells were infected with chlamydiae and then exposed to dilute concentrations of C31G which did not alter epithelial cell viability, chlamydial infectivity was also markedly reduced. C31G gained access to the developing chlamydial inclusion causing damage to or destruction of metabolically active reticulate bodies as well as apparent alteration of the inclusion membrane, which resulted in premature escape of chlamydial antigen to the infected epithelial surface. These studies show that the broad-spectrum antiviral and antibacterial microbicide C31G also has antichlamydial activity.

Anti-Bacterial Agents↗
PubMed1996

Assessment of the anti-microbial agent C31G as a spermicide: comparison with nonoxynol-9.

The broad-spectrum anti-microbial agent, C31G, containing an equimolar mixture of n-dodecyl-dimethylamine-N-oxide (C12-N-O) and N-(n-dodecyl), N-dimethyl-glycine (C12-betaine), was tested for spermicidal activity in comparison with the currently used spermicide, nonoxynol-9 (N-9). The rate of sperm cell permeabilization by the spermicides, as assayed with the fluorescent probe, TO-PRO-1, increased as the cube of the C31G concentration, while the rate increase was linear with N-9 concentration. At 0.04%, the rate of sperm cell permeabilization with both spermicides is at the limit of rapid measurement. C31G diffuses through cervical mucus at a more rapid rate than does N-9. C31G has long been known to aid wound healing and reduce inflammation, whereas N-9 has been reported to induce vaginal irritation. C31G would, thus, seem to have the spermicidal efficacy, the broad range of anti-microbial activity, and the lack of inflammatory activity that is sought in the ideal vaginal spermicide.

Adult↗
PubMed1994

Molecules of Streptococcus gordonii that bind to Porphyromonas gingivalis.

Interbacterial binding is considered an important colonization mechanism for many of the organisms that inhabit dental plaque. Porphyromonas gingivalis, a periodontal pathogen, can adhere to species that comprise early plaque, such as Streptococcus gordonii. In this study, the molecules of S. gordonii G9B that mediate binding to P. gingivalis were investigated. Biotinylated surface molecules of S. gordonii were extracted and mixed with P. gingivalis cells. Interactive streptococcal components were identified by SDS-PAGE of the P. gingivalis cells followed by electroblotting, and visualization of the adsorbed streptococcal molecules with streptavidin-alkaline phosphatase. S. gordonii molecules of 45 kDa and a doublet of 62/60 kDa were observed to bind to P. gingivalis. Polyclonal antibodies raised to the 62/60 kDa proteins inhibited the binding interaction. These antibodies demonstrated an antigenic relationship between the 62/60 kDa molecules and the 45 kDa protein. Both molecules were also antigenically related to, and may be breakdown products of, a larger molecule of 170 kDa which is antigenically related to the P1 antigen of S. mutans. Cloning and expression in Enterococcus faecalis of the gene for the P1-like molecule from S. gordonii M5 resulted in a phenotype that expressed the 62/60 kDa and 45 kDa antigens and was capable of binding to P. gingivalis. These results suggest that a P1-like molecule in S. gordonii is involved in adherence to P. gingivalis. Processing of the P1-like molecule into smaller fragments of 62/60 kDa and 45 kDa may be required for binding activity.

Animals↗
PubMed1994

Calcium-binding properties of SSP-5, the Streptococcus gordonii M5 receptor for salivary agglutinin.

Streptococcus gordonii M5 expresses a lectin on its surface (SSP-5) which binds to human salivary agglutinin (SAG). This interaction requires sialic acid residues of SAG and divalent cations and may mediate the colonization of oral tissues by this organism. In this report, we show that the binding of SAG to SSP-5 requires calcium and that SSP-5 is a high-affinity calcium-binding protein. SAG-mediated aggregation of S. gordonii M5 was inhibited by 1 mM EDTA, and the restoration of aggregation occurred only upon the readdition of calcium. To ascertain the level at which calcium exerts its effects, the calcium-binding properties of SSP-5 were evaluated by using a 45Ca binding assay. In addition, a kinetic analysis of calcium binding was carried out by using fura2, a fluorescent calcium-binding dye. These analyses showed that SSP-5 is a high-affinity calcium-binding protein that binds 1 mol of calcium per mol of protein and has a dissociation constant of 0.45 +/- 0.2 microM. The calcium-binding capacity of SSP-5 was also calculated independently to be 1.0 +/- 0.2 mol of Ca per mol of SSP-5 by column chromatography on Sephadex G-25 equilibrated with 10 microM 45Ca. To localize the calcium binding site of SSP-5, a series of C-terminal deletion mutants were expressed in Escherichia coli and evaluated for calcium-binding activity. Deletion of the 250 C-terminal residues of SSP-5 had little effect on calcium binding. However, deletion of residues 1168 to 1250 resulted in the loss of calcium-binding activity, suggesting that this region is important for calcium binding by SSP-5.

Adhesins, Bacterial↗
PubMed1994

Role of the cytoskeleton in cell-to-cell transmission of human immunodeficiency virus.

We previously observed that when human immunodeficiency virus (HIV)-infected T lymphocytes are added to epithelial cells, they adhere, polarize, and secrete virions unidirectionally onto the epithelium. Epithelial cells subsequently take up virus and become productively infected. We report here that colchicine treatment of T-lymphocyte suspensions induced lymphocyte polarization, redistribution of F-actin into a pseudopod, and secretion of HIV from the pseudopod. Immobilization of T lymphocytes on negatively charged plastic also caused redistribution of F-actin and unidirectional secretion of HIV onto the plastic. As neither colchicine nor adhesion caused an increase in HIV secretion, they apparently act by focusing secretion to the tip of the pseudopod. We speculate that adhesion-induced polar secretion of HIV, from activated mononuclear cells onto epithelia, is a cytoskeleton-mediated process which may be involved in HIV transmission in vivo.

Actins↗
PubMed1993

Human submandibular saliva aggregates HIV.

Incubation of HIV with human whole, parotid, or submandibular saliva leads to a decrease in viral infectivity in Sup-T1 cells. The effect is most pronounced with submandibular saliva. Inhibition is seen within 2 min, and increases with time. There is little inhibition seen after incubation of saliva with HSV, and no effect with adenovirus, suggested that there is some viral specificity. Electron microscopic studies revealed that HIV-saliva aggregates are trapped in 0.45-microns pore size nitrocellulose filters. If these inhibitory effects are manifest in vivo, this could account for the low level of virus detected in oral secretions.

Adenoviridae↗
PubMed1993

Localization of salivary proteins in granules of human parotid and submandibular acinar cells.

Electron microscopic immunocytochemistry was used to investigate the distribution of four secretory proteins within the granules of human parotid and submandibular gland acinar cells. The results demonstrate that the distribution of the individual proteins differs significantly. Amylase is concentrated within the electron-dense regions of the granules, whereas agglutinin is present mainly in the regions of lower density. In contrast, the proline-rich proteins and histatins are distributed throughout the content of the granules and do not appear to be concentrated in either the high- or low-density regions. The results also suggest that different mechanisms, such as self-aggregation or ionic complexes with metal ions and charged macromolecules, may be involved in the packaging of individual proteins within the same granule. Further, they also show that among the salivary glands of various species, the intragranular distribution of similar proteins (e.g., amylase and proline-rich proteins) may differ.

Agglutinins↗
PubMed1993

HIV in the oral cavity: virus, viral inhibitory activity, and antiviral antibodies: a review.

Although it is generally assumed that HIV transmission does not occur through casual oral contact, persistent reports in the literature and the well-documented case of the Florida dentist (Ou et al., 1992) have served to elevate concerns and interest about the possibility of oral transmission of HIV. The literature suggests that: (1) the presence of infectious virus in the oral cavity is an uncommon event; (2) PCR data indicate that HIV sequences may be present in the oral cavity at reasonably high frequency--further studies are warranted; (3) saliva appears to contain potent anti-HIV activity that may be responsible for the low oral virus titer; and (4) oral secretions are a reliable source for monitoring anti-HIV antibodies. It is clear that the oral cavity will remain a focus for HIV research, in terms of both viral transmission/pathogenesis and for noninvasive diagnosis of the HIV-positive individual.

Antiviral Agents↗
PubMed1992

Differential distribution of a carbohydrate epitope (Y) on human salivary gland cell membranes.

Monoclonal antibody 303 (mAb 303) reacts with the high molecular weight agglutinin present in human saliva. Its reactivity is periodate sensitive, and it has been shown to recognize the Y epitope. Immunogold labeling of thin sections of human parotid and submandibular glands with mAb 303 showed reactivity in secretory granules of serous acinar, intercalated and striated duct cells (Takano et al., 1988). We now report that the apical and basolateral membranes of salivary acinar and duct cells are labeled by mAb 303, but not myoepithelial cells, endothelial cells and other mesenchymal cells. Gold particles were confined to acinar and duct cell membranes even when myoepithelial cells were directly adjacent, suggesting that the epitope resides on a membrane glycoprotein and that the label does not represent secreted agglutinin bound to the cell surface. Although myoepithelial cells are thought to differentiate from epithelial stem cells, the present results indicate that substantial compositional differences exist between the membranes of myoepithelial cells and other salivary parenchymal cells. Earlier studies also showed that mAb 303 labels normal pancreatic acinar cells and certain salivary (pleomorphic adenoma) and mammary (lactating adenoma) tumors (Bogert et al., 1988). This antibody thus may be a useful reagent for characterizing the origin of exocrine gland-derived cell cultures and neoplastic cells. Further, localization studies may provide insight into the role of the Lewis blood group-related epitope in secretory cells.

Antibodies, Monoclonal↗
PubMed1991

Differential distribution of salivary agglutinin and amylase in the Golgi apparatus and secretory granules of human salivary gland acinar cells.

The secretory granules of salivary glands often display complex internal substructures, yet little is known of the molecular organization of their contents or the mechanisms involved in packaging of the secretory proteins. We used post-embedding immunogold labeling with antibodies to two secretory proteins, agglutinin and alpha-amylase, to determine their distribution in the Golgi apparatus and secretory granules of the human submandibular gland acinar cells. With monoclonal antibodies specific for carbohydrate epitopes of the agglutinin, reactivity was found in the trans Golgi saccules, trans Golgi network, and immature and mature secretory granules. In the granules, labeling was seen in regions of low and medium electron density, but not in the dense cores. Reactivity seen on the apical and basolateral membranes of acinar and duct cells was attributed to a shared epitope on a membrane glycoprotein. Labeling with a polyclonal antibody to amylase was found in the Golgi saccules, immature and mature secretory granules, but not in the trans Golgi network. In the granules, amylase was present in the dense cores and in areas of medium density, but not in the regions of low density. These results indicate that these two proteins are distributed differently within the secretory granules, and suggest that they follow separate pathways between the Golgi apparatus and forming secretory granules. Small vesicles and tubular structures that labeled only with the antibodies to the agglutinin were observed on both faces of the Golgi apparatus and in the vicinity of the cell membrane. These structures may represent constitutive secretion vesicles involved in transport of the putative membrane glycoprotein to the cell membrane.

Agglutinins↗
PubMed1991

Salivary-agglutinin-mediated adherence of Streptococcus mutans to early plaque bacteria.

Interspecies binding is important in the colonization of the oral cavity by bacteria. Streptococcus mutans can adhere to other plaque bacteria, such as Streptococcus sanguis and Actinomyces viscosus, and this adherence is enhanced by saliva. The salivary and bacterial molecules that mediate this interaction were investigated. Salivary agglutinin, a mucinlike glycoprotein known to mediate the aggregation of many oral streptococci in vitro, was found to mediate the adherence of S. mutans to S. sanguis or A. viscosus. Adherence of S. mutans to saliva- or agglutinin-coated S. sanguis and A. viscosus was inhibited by antibodies to the bacterial agglutinin receptor. Expression of the S. sanguis receptor (SSP-5) gene in Enterococcus faecalis increased adhesion of this organism to saliva- or agglutinin-coated S. sanguis and A. viscosus. This interaction could be inhibited by antibodies to the agglutinin receptor. The results suggest that salivary agglutinin can promote adherence of S. mutans to S. sanguis and A. viscosus through interactions with the agglutinin receptor on S. mutans.

Agglutinins↗
PubMed1990

Streptococcal-host interactions. Structural and functional analysis of a Streptococcus sanguis receptor for a human salivary glycoprotein.

Colonization of oral tissues by Streptococcus sanguis may be influenced by a mucin-like salivary glycoprotein (SAG) through a calcium-dependent interaction with a specific bacterial receptor. We report the nucleotide and deduced amino acid sequence of the S. sanguis receptor (SSP-5) and show that this protein may bind sialic acid residues of SAG. The SSP-5 protein contains three unique structural domains, two of which consist of repetitive amino acid sequences. The N-terminal domain is comprised of four tandem copies of an 82-residue repeat which exhibits homology to M protein of Streptococcus pyogenes. This region is highly charged and predicted to be alpha-helical. A second hydrophilic repetitive domain consists of three copies of a 39-amino acid sequence containing 30% proline flanked by nonrepetitive proline-rich sequence. The third domain consists of 48% proline and resides near the C terminus of the protein. Secondary structure analysis of the SSP-5 sequence also identified four potential helix-turn-helix motifs that resembled E-F hand calcium binding domains. The SSP-5 protein is highly homologous to a surface antigen expressed by the mutans streptococci and the domain structure of SSP-5 is conserved within this family of proteins. The interactions of SSP-5 and of intact S. sanguis with SAG were inhibited by neuraminidase digestion of the salivary glycoprotein and by simple sugars containing sialic acid, suggesting that sialic acid is the primary ligand involved in the binding reaction.

Amino Acid Sequence↗
PubMed1990

Comparison of Streptococcus mutans and Streptococcus sanguis receptors for human salivary agglutinin.

Oral streptococci vary in their susceptibility to salivary agglutinin-mediated aggregation. To understand the molecular basis of this specificity, the structure and function of receptors for agglutinin from Streptococcus mutans KPSK2 (MSL-1) and Streptococcus sanguis M5 (SSP-5) were compared. Immunological screening of an S. mutans KPSK2 genomic DNA library yielded two identical clones expressing a streptococcal protein that co-migrated with a 220 kDa peptide in SDS extracts from this organism. This protein inhibited agglutinin-mediated aggregation of S. mutans KPSK2 in a dose-dependent manner. The MSL-1 gene is homologous to the S. mutans SpaP and pac genes although single base substitutions alter several amino acids. MSL-1 is also similar to the agglutinin receptor (SSP-5) cloned from S. sanguis M5. All three proteins, MSL-1, P1, and SSP-5 share at least one epitope since monoclonal and polyclonal anti-SSP-5 antibodies react with both MSL-1 and P1. However, other monoclonal antibodies are specific for SSP-5 and appear to react with a peptide domain exhibiting little homology to MSL-1 or P1. Sugar inhibition studies showed that agglutinin-mediated aggregation of S. mutans KPSK2 was most potently inhibited by fucose and lactose. Sialic acid, a potent inhibitor of S. sanguis aggregation, had no effect on the interaction of agglutinin with S. mutans KPSK2. These results suggest that while the MSL-1 and SSP-5 proteins are genetically and immunologically related, their specificity for binding sites on agglutinin differs.

Adhesins, Bacterial↗
PubMed1990

Clinical study of a C31G containing mouthrinse: effect on salivary microorganisms.

In vitro studies have demonstrated the antiplaque properties of C31G, a potent broad spectrum antimicrobial agent consisting of an equimolar mixture of alkyl dimethyl glycine and alkyl dimethyl amine oxide, buffered with citric acid. In this initial clinical study, C31G at concentrations of 0.05%, 0.1%, 0.2% and 0.5%. Listerine, and placebo were tested in a complete crossover design. Twelve subjects were evaluated, with a minimum of 2 days between treatments. Parameters monitored were salivary bacterial counts and saliva glycolysis. The 0.5% and 0.2% C31G mouthrinses significantly reduced total bacterial counts in saliva samples obtained up to and including three hours after rinsing, compared with counts obtained prerinsing or after placebo rinsing. Both 0.5%, and 0.2% C31G significantly inhibited glycolysis of salivary bacteria for up to 6 hours postrinsing, compared with pH values obtained prerinsing. 0.1% and 0.05% C31G exhibited little or no effect in either assay. Listerine showed a significant reduction in bacterial counts for up to 1 hour postrinsing, compared with prerinse counts, but the effect was less sustained. Listerine showed no significant inhibition of glycolysis at any time point. No tooth staining or altered taste sensation was noted with either product.

Adult↗