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T J Speaker

Publications and source records attributed to T J Speaker.

At least 19 recordsLinked to original sources
PubMed2000

A quantitative luminescence assay for measuring cell uptake of aqueous-based microcapsules in vitro.

We recently developed a system of microencapsulation consisting of aqueous-based polymers (e.g. alginate) and aqueous amines (e.g. spermine). We found that microencapsulation enhanced virus-specific protective immune responses. In addition, we found that microencapsulation may enhance virus-specific immune responses by selecting for antigen-presenting cells (APC) that are more efficient at processing and presenting viral antigens than those involved after natural infection. To determine the intracellular trafficking patterns and fate of microcapsules within APC, we developed a luminescence assay that permits the determination of specific quantities of proteins introduced into cells by microcapsules. We found that the time-dependent uptake of horseradish peroxidase (HRP)-labeled microcapsules was accurately detected in lysates of peritoneal exudate cells using luminol. The amplitude of HRP-catalyzed chemiluminescence in cell lysates correlated with the capture efficiency and retention kinetics of HRP in three different microcapsule preparations. HRP was most efficiently captured and retained by linking biotinylated HRP to microcapsulses chemically modified at the amine moiety with egg avidin. This preparation yielded more accurate and sensitive quantitation of HRP contained within cells than preparations capturing HRP or HRP-conjugated goat antibody into the microcapsular matrix by ionic interactions.

Animals↗
PubMed2000

Water-based microsphere delivery system for proteins.

This paper describes formulation of a model protein, horseradish peroxidase (HRP), in a water based microcapsule delivery system and demonstrates the utility of this delivery system for proteins. Aqueous solutions (1 mg/mL) of the enzyme were separately blended with aqueous solutions of the neutral sodium salt of the anionic polymer iota carrageenan (0.6 mM in repeat unit). These blends were instilled as uniform microdroplets into aqueous solutions of a series of eleven mono-, di-, or oligo-amines (as neutral hydrochloride or acetate salts). Essentially instantaneous salt exchange interaction of the sodium salt of anionic polymer with amine hydrochloride resulted in formation of microparticles of amine/polymer complex. The enzyme was captured in the resulting capsules. The particles were washed by repeated centrifugation and resuspension in water and their particle size distribution was determined. HRP in washed pelleted microspheres was analyzed for fragmentation/aggregation by SDS-PAGE and size exclusion chromatography, for unfolding by fluorescence spectroscopy, and for specific enzymatic activity, capture efficiency and release studies by absorbance spectroscopy. Dependent on amine employed, capture efficiencies ranged from 1 to 72%. Encapsulation produced no adverse effect on protein size as no molecular fragments or aggregates were visible below or above 44 kDa. The tryptophan fluorescence spectrum of the protein did not change after encapsulation indicating no conformational change in tertiary structure. There was an apparent substrate diffusion related reduction in activity of encapsulated HRP, but almost 100% of activity was recovered on lysis of the capsules. It is concluded that water based charged film encapsulation used as a drug delivery system for proteins does not alter structural conformation or specific activity of the model protein tested and provides protein release at a constant rate.

Carrageenan↗
PubMed1998

Effect of water-based microencapsulation on protection against EDIM rotavirus challenge in mice.

We determined the capacity of microcapsules formed by the combination of sodium alginate, an aqueous anionic polymer, and spermine hydrochloride, an aqueous cationic amine, to enhance protection against rotavirus challenge in mice. Adult BALB/c mice were orally inoculated with either free or microencapsulated rotavirus (simian rotavirus strain RRV) and challenged 6 or 16 weeks later with murine rotavirus strain EDIM. Virus-specific humoral immune responses were determined at the time of challenge and 4 days after challenge by intestinal fragment culture. We found that spermine-alginate microcapsules enhanced protection against challenge 16 weeks after immunization but not 6 weeks after immunization. Quantities of virus-specific immunoglobulin A produced by small intestinal lamina propria lymphocytes were correlated with the degree of protection against challenge afforded by spermine-alginate microcapsules. Possible mechanisms by which microcapsules enhance protection against rotavirus challenge are discussed.

Alginates↗
PubMed1997

Effect of microencapsulation on immunogenicity of a bovine herpes virus glycoprotein and inactivated influenza virus in mice.

We previously found that aqueous-based spermine-alginate or spermine-chondroitin sulfate microcapsules enhanced rotavirus-specific humoral immune responses after intramuscular inoculation of mice. To extend our observations with whole, infectious rotavirus to vaccine strategies which include inactivated virus and purified proteins, we determined the capacity of aqueous-based microcapsules to enhance virus-specific immune responses to bovine herpes virus type 1 glycoprotein D (BHV-1-gD) or ether-treated influenza virus. We found that spermine-alginate microcapsules decreased the quantity of BHV-1-gD necessary to induce protein-specific antibodies about 5000-fold. However, spermine-alginate microcapsules did not enhance influenza virus-specific antibody responses. Microcapsules composed of spermine-chondroitin sulfate did not enhance either BHV-1-gD or influenza virus-specific immune responses. Possible mechanisms of enhancement of virus-specific antibody responses by microencapsulation are discussed.

Alginates↗
PubMed1997

Aqueous-based microcapsules are detected primarily in gut-associated dendritic cells after oral inoculation of mice.

We previously found that aqueous-based microencapsulation enhanced virus-specific humoral immune responses after oral inoculation of mice. However, the mechanism by which microencapsulation enhances immunogenicity remains unclear. We found that spermine-alginate microcapsules were detected primarily in gut-associated dendritic cells (i.e. CD11c/CD18+, Ia+, CD11b-, CD45R-) after oral inoculation of adult mice. Microencapsulation may enhance immunogenicity by involving antigen presenting cells which are more efficient than those recruited during natural infection.

Administration, Oral↗
PubMed1997

Orally administered microencapsulated reovirus can bypass suckled, neutralizing maternal antibody that inhibits active immunization of neonates.

Purified reovirus serotype 1, encapsulated in biodegradable aqueous microcapsules, was found to bypass maternal antibody passively transferred by suckling to neonates. Genetically identical, immunocompetent F1 scid/+ mice were generated by the reciprocal crosses of C.B17 scid/scid and normal congenic +/+ adult mice. The immunocompetent +/+ dams were either orally infected with reovirus prior to mating or not. Thus, these immunocompetent F1 pups developed either in the absence or in presence of passively transferred maternal immunity. The F1 mice were orally immunized on day 10 with either live virus, microencapsulated reovirus, or empty microcapsules plus live virus. The immune responses were assessed in the neonatal gut-associated lymphoid tissues (GALT). Examination of reovirus specific immunoglobulin A in the serum and GALT, taken on days 7, 14, and 21 postimmunization, clearly demonstrated that microencapsulated reovirus could bypass the normal effect of maternal antibodies, passively acquired by suckling, to inhibit active priming of neonates by oral route. These observations seem relevant to the development of efficacious oral vaccines that also allow passive, protective immunity via suckled maternal antibodies while permitting active oral immunization of neonates.

Administration, Oral↗
PubMed1996

Aqueous-based microencapsulation enhances virus-specific humoral immune responses in mice after parenteral inoculation.

Vaccines are commonly administered by the parenteral route. Therefore, adjuvant strategies which include parenteral immunization may improve the efficacy of a number of current vaccines. The capacity of aqueous-based microencapsulation to enhance virus-specific IgG responses in mice inoculated intramuscularly with small quantities of antigen was evaluated. Mice were inoculated with either 10(4), 10(3), or 10(2) p.f.u. of microencapsulated rotavirus (bovine strain WC3), placebo microcapsules plus free virus, or virus alone. Mice were subsequently bled 1, 2, 4, 6, and 9 months after inoculation. Microencapsulation of rotavirus enhanced virus-specific humoral immune responses. In addition, virus-containing microcapsules composed of spermine-chondroitin sulfate induced levels of virus-specific antibodies greater than those found after inoculation with virus-containing microcapsules composed of spermine-alginate. Mechanisms by which microencapsulation may enhance virus-specific humoral immunity are discussed.

Animals↗
PubMed1995

Enhancement by microencapsulation of rotavirus-specific intestinal immune responses in mice assessed by enzyme-linked immunospot assay and intestinal fragment culture.

The capacity of microencapsulation to enhance the humoral immune response to rotavirus in the gut-associated lymphoid tissue (GALT) of mice was determined by using a system of microencapsulation based on the ionic linkage of aqueous anionic polymers and an aqueous amine. Inoculation of mice with microencapsulated rotavirus enhanced the frequencies of virus-specific IgA-secreting cells in the lamina propria as well as the quantities of virus-specific IgA produced in GALT. In addition, an enhanced virus-specific immune response was associated with enhanced production of presumably polyclonal, non-rotavirus-specific antibodies in GALT. The mechanism by which microencapsulation enhances the humoral immune response remains to be determined.

Alginates↗
PubMed1995

Oral inoculation of mice with low doses of microencapsulated, noninfectious rotavirus induces virus-specific antibodies in gut-associated lymphoid tissue.

The capacity of an aqueous-based system of microencapsulation to enhance virus-specific humoral immune responses was evaluated in mice orally inoculated with noninfectious rotavirus (simian rotavirus strain RRV). Mice were orally inoculated with 1.75 or 0.35 microgram of inactivated RRV (iRRV) or microencapsulated iRRV. Sera, intestinal contents, and organ cultures of gut-associated lymphoid tissues (GALT) were tested for the presence of rotavirus-specific antibodies. Virus-specific IgA was produced by small intestine lamina propria lymphocytes in animals inoculated with 1.75 or 0.35 microgram of microencapsulated virus, but not in mice inoculated with unencapsulated virus. Virus-specific IgA in sera and intestinal contents were not predictive of intestinal organ culture responses. Microencapsulation may be an efficient way of inducing virus-specific immune responses in GALT after oral inoculation with small quantities of viral antigen. In addition, delayed release of virus from microcapsules may obviate the need for booster immunizations.

Administration, Oral↗
PubMed1994

Enhancement of rotavirus immunogenicity by microencapsulation.

It was determined whether microencapsulation of rotavirus enhanced virus-specific immunity in mice. Combinations of several water-soluble anionic polymers and amines were tested for their capacity to form microcapsules which were stable in the presence of simulated gastric acid. Using the combinations of sodium alginate and spermine hydrochloride or sodium chondroitin sulfate and spermine hydrochloride we found that microcapsules (1) captured infectious rotavirus, (2) penetrated into the persisted in gut-associated lymphoid tissue (GALT) after oral inoculation, (3) delivered rotavirus antigen to GALT at levels greater than those detected after oral inoculation with free virus, and (4) enhanced the virus-specific humoral immune response after oral or parenteral immunization.

Amines↗
PubMed1980

Substituent constants of azulene.

The ionization constants in water for six 3-substituted azuloic acids were determined spectrophotometrically. Conversion of these physical constants to their pKa values allowed a set of Hammett-type sigma values for the substituents on these acids to be calculated. Determination of partition coefficients for nine 1-substituted azulenes allowed Hansch-type pi values to be determined, using azulene as the model compound.

Azulenes↗
PubMed1975

Azulene analogs of pharmacological agents III: acute toxicity and local anesthetic activity of azulylamides and azulenecarboxamides.

This paper describes the acute toxicity of two azulylamides and six azulenecarboxamides and the nerve conduction-inhibiting properties of some of these compounds. The azulene derivatives are compared to their benzenoid prototypes, lidocaine and procainamide, as part of a continuing investigation of the biological properties of nonbenzenoid aromatic compounds.

Acetamides↗