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Biomedical subjects

M W Russell

Publications and source records attributed to M W Russell.

At least 55 records · Page 3Linked to original sources
PubMed1997

Establishment of a Streptococcus pneumoniae nasopharyngeal colonization model in adult mice.

Human nasopharyngeal carriage of Streptococcus pneumoniae constitutes the major natural reservoir of pneumococci and is thought to be the prelude to virtually all pneumococcal disease. If carriage could be greatly reduced, pneumococcal transmission and disease could be largely eliminated. To facilitate the studies of mechanisms important in carriage and to identify immunogens that can elicit protection against carriage, we characterized an adult mouse model of nasopharyngeal carriage. Non-anaesthetized mice were inoculated intranasally with pneumococci in 10 microl of fluid. Nasopharyngeal carriage was observed with strains of capsular types 3, 4, 6A, 6B, 14, 19, and 23. Carriage was stable over time, and the numbers of pneumococci carried were relatively independent of inoculation dose; findings which indicate that the recovery of pneumococci from 1 day to 2 weeks post inoculation was dependent on colonization, rather than just temporary contamination. To ensure carriage in the largest percentage of mice, without causing sepsis or death, inoculations of 10(7) colony forming units (cfu) should be used. In this model, carriage was generally observed without concomitant bacteremia or sepsis and carriage was observed even with strains that were avirulent when injected i.v. The model should be useful for the identification of protection-eliciting antigens, since intranasal immunization with heat-killed pneumococci or lysates of pneumococci protected against carriage.

Animals↗
PubMed1997

Nasal lymphoid tissue, intranasal immunization, and compartmentalization of the common mucosal immune system.

Mucosal application of vaccines with an appropriate adjuvant can induce immune responses at both systemic and mucosal sites, and therefore may prevent not only infectious disease, but also colonization of mucosal surfaces. Intranasal is more effective than intragastric immunization at generating earlier and stronger mucosal immune response. Nasal lymphoid tissue (NALT) and its local draining lymph nodes may retain long-term immune memory. IgA isotype switching, and the differentiation and maturation of IgA antibody-secreting cells (ASC) may occur before these cells migrate out of NALT, whereas IgG ASC responses require passage of the cells through draining lymph nodes of the NALT. Knowledge of whether immune memory cells can recirculate to and reside in the inductive sites other than their origin after encountering antigen will be helpful for understanding the compartmentalization of the common mucosal immune system as well as for determining the best route for delivering a mucosal vaccine against a particular pathogen.

Administration, Intranasal↗
PubMed1997

Nasal lymphoid tissue (NALT) as a mucosal immune inductive site.

Intranasal (i.n.) immunization is a very effective route for inducing mucosal immunity, but the cellular mechanism responsible for regulating and disseminating these responses is not fully understood. The authors studied the role of nasal lymphoid tissue (NALT) as a mucosal inductive site by using highly purified NALT cells obtained by a new method of mechanical isolation. The NALT cells, like Peyer's patch (PP) cells, were smaller in size and less granular than lymphocytes from salivary glands (SG) and small intestinal lamina propria (LP). The NALT cells isolated from i.n. immunized mice contained antigen-specific antibody-secreting cells (ASC) predominantly of immunoglobulin (Ig)A isotype, similar to those also recovered from salivary glands in a time course study. However, the higher proportion of smaller sized sports formed by NALT cells in ELISPOT assays suggested that these cells were less differentiated precursors of those found in salivary glands. This was supported by the fact that after i.n. immunization, IgA ASC appeared in NALT, as well as in mucosal effector sites SG and LP, but none or very few in another mucosal inductive site, PP. In contrast, after intragastric (i.g.) immunization, IgA ASC were detected in PP, along with the SG and LP, but none or very few in NALT. Furthermore, after i.n. immunization, lymphocytes from NALT but not PP proliferated in response to the specific antigen in culture. These findings imply that NALT served as an inductive site for IgA antibody responses at mucosal effector sites.

Administration, Intranasal↗
PubMed1997

Intranasal immunization of mice with PspA (pneumococcal surface protein A) can prevent intranasal carriage, pulmonary infection, and sepsis with Streptococcus pneumoniae.

Many pathogens, including Streptococcus pneumoniae, are carried asymptomatically on the nasopharyngeal mucosa and spread among individuals by close contact. Clinical disease results when pneumococci escape from the mucosa and invade sterile sites. Although systemic immunity can prevent invasive disease, control of person-to-person spread is probably dependent on immunity acting at the mucosal surface. Intranasal immunization of mice with PspA (pneumococcal surface protein A) or a capsular 6B polysaccharide-tetanus toxoid conjugate induced mucosal and systemic antibody responses and provided long-lasting protection against carriage of S. pneumoniae. Resistance to carriage was dependent on mucosal rather than systemic immunity and was effective against heterologous strains of heterologous PspA types. Intranasal immunization with PspA also protected against systemic infection following intravenous, intratracheal, and intraperitoneal challenge.

Animals↗
PubMed1997

Development of antibody-secreting cells and antigen-specific T cells in cervical lymph nodes after intranasal immunization.

Intranasal (i.n.) immunization with bacterial protein antigens coupled to cholera toxin B subunit (CTB) effectively induces mucosal, especially salivary immunoglobulin A (IgA), and nonmucosal antibody responses in mice. To examine the regional distribution of antigen-specific B and T cells after i.n. immunization, antibody-secreting cells and antigen-responsive T cells in cervical lymph nodes (CLN) were compared with those found after intraoral or subcutaneous (in the neck) administration of the same antigen and with T cells found in mesenteric lymph nodes (MLN) and spleen after intragastric immunization. The i.n. immunization induced predominantly IgA antibody-secreting cells in salivary glands and IgA and IgG antibody-secreting cells in the superficial and central CLN; these responses were quantitatively enhanced if the antigen was coupled to CTB. Intraoral immunization also induced IgA and IgG antibody-secreting cells in the superficial and central CLN, but only if intact cholera toxin was included as an adjuvant. In contrast, subcutaneous (neck) immunization induced IgG antibody-secreting cells mainly in the draining facial lymph nodes. CLN cell populations resembled those of MLN, except that CLN lymphocytes had higher proportions of T cells and lower proportions of B cells and a slightly higher CD4+/CD8+ ratio among T cells than the MLN lymphocytes did. T cells that proliferated in response to antigen in vitro were found especially in central CLN 2 days after i.n. immunization and persisted for up to 6 months, whereas after intragastric immunization, responsive T cells were not found in the MLN for up to 14 days. After culture with antigen in vitro, T cells from the superficial CLN of i.n. immunized mice secreted both gamma interferon and interleukin-4. Therefore, after i.n. immunization, superficial and central CLN represent sites of regional lymphocyte development, and the central CLN in particular appear to be sites where memory T cells persist.

Adjuvants, Immunologic↗
PubMed1997

Oral immunization with the saliva-binding region of Streptococcus mutans AgI/II genetically coupled to the cholera toxin B subunit elicits T-helper-cell responses in gut-associated lymphoid tissues.

Mice immunized intragastrically (i.g.) with a genetically constructed chimeric protein consisting of the saliva-binding region (SBR) of Streptococcus mutans AgI/II coupled to cholera toxin (CT) A2 and B subunits (CTA2/B) develop serum immunoglobulin G (IgG) and mucosal IgA antibody responses against AgI/II that are enhanced by the coadministration of CT as an adjuvant. To investigate the development of antigen-specific T cells in the gut-associated lymphoid tissues, mice were immunized i.g. with SBR, SBR-CTA2/B, or SBR-CTA2/B plus CT. AgI/II-specific T cells in Peyer's patches (PP), mesenteric lymph nodes (MLN), and spleen were assayed by lymphoproliferation and flow cytometry for the expression of T-cell surface markers, and cytokine mRNA expression was evaluated by reverse transcription-PCR. T-cell responses were consistent with antibody responses but were detectable after the first immunization. Proliferative responses of PP and MLN cells upon stimulation with AgI/II in vitro were low and delayed in mice given SBR alone, and these cells displayed a mixed type 1 and 2 (or Th0) pattern of cytokine expression. Immunization with SBR-CTA2/B resulted in greater AgI/II-specific proliferative responses in PP cells and an increase in the proportion of CD4+ T cells. Coadministration of CT with SBR-CTA2/B led to greater proliferative responses especially in the MLN cells, which then showed an increase in CD4+ cells. Immunization with SBR-CTA2/B (with or without CT) skewed the cytokine expression pattern in PP and MLN cells toward Th2. The results indicate that T helper cells were induced in gut-associated lymphoid tissues by i.g. immunization with SBR-CTA2/B, concomitantly with and prior to the appearance of circulating and mucosal antibodies.

Administration, Oral↗
PubMed1997

Mucosal immunogenicity of a recombinant Salmonella typhimurium-cloned heterologous antigen in the absence or presence of coexpressed cholera toxin A2 and B subunits.

An avirulent Salmonella typhimurium vaccine strain expressing a streptococcal protein adhesin and a similar clone which produces the same streptococcal antigen linked to the cholera toxin (CT) A2 and B subunits (CTA2/B) were compared for the ability to induce antibody responses to the expressed heterologous antigen after oral or intranasal immunization of mice. Expression of cloned immunogens in these systems is temperature regulated, being optimal at 37 degrees C, and the two clones under comparison were shown to produce similar levels of the streptococcal antigen. Both clones were found to stimulate high levels of serum immunoglobulin G (IgG) and mucosal IgA antibodies to the cloned immunogen. A consistent trend was observed toward higher mucosal IgA but lower serum IgG responses in the case of the S. typhimurium vector that coexpressed CTA2/B, a potential mucosal adjuvant, regardless of the route of administration. Also noteworthy was the capacity of these antigen delivery systems to induce anamnestic systemic and secretory responses to the cloned immunogen 15 weeks after the primary immunization, despite preexisting immunity to the Salmonella vectors. These antibody responses were sustained for at least 7 months following the booster immunization, at which time the secretory IgA antibody levels were significantly higher in mice given the Salmonella clone that coexpressed CTA2/B. Although the serum IgG response against the Salmonella vector was characterized by a high IgG2a/IgG1 ratio (indicative of the T helper type 1 [Th1]/Th2 profile), a mixed IgG1 and IgG2a pattern was observed for the carried heterologous antigen, which displayed a dominant IgG1 response when administered as a purified immunogen. Our findings indicate that the recombinant streptococcal antigen and CTA2/B are strong immunogens when expressed by the antigen delivery system used in this study and suggest that CTA2/B may have an additional immunoenhancing activity in the mucosal compartment besides its ability to target antigen uptake into the mucosal inductive sites. CTA2/B may thus be useful as an S. typhimurium-cloned adjuvant for coexpressed protein antigens.

Animals↗
PubMed1997

Routes of immunization and antigen delivery systems for optimal mucosal immune responses in humans.

Numerous experiments performed in humans and animals have revealed that stimulation of mucosal lymphoid inductive sites such as intestinal Peyer's patches results in parallel immune responses manifested by the appearance of S-IgA antibodies in the external secretions of remote glands. However, recent experiments suggest that inductive sites associated with the upper respiratory tract, rectum, and perhaps genital tract may also function as sources of lymphoid cells that populate, with some selectivity, certain remote mucosal effector sites. Furthermore, antigen-specific IgA antibodies can be induced in certain secretions (e.g., female genital tract) not only by immunization in the vicinity of corresponding mucosal tissues (e.g., vagina and rectum) but also by oral and especially intranasal immunization. The ineffectiveness of simple delivery of soluble antigens to mucosal membranes for immunization has stimulated extensive studies of strategies for effective delivery systems that would (a) increase the antigen absorption, (b) prevent its degradation, and (c) skew the outcome of immunization to a desired goal (protective response to infectious diseases vs. tolerance; B vs. T cell responses; mucosal vs. systemic). The induction of immune responses at a desired mucosal site can be accentuated with the use of a suitable antigen-delivery system including relevant bacterial or viral vectors, edible transgenic plants expressing microbial antigens, incorporation of antigens in biodegradable microspheres or liposomes, and linkage or coadministration of antigens with cholera toxin B subunit. However, only a few antigen-delivery systems extensively used in animal experimentation have been evaluated for their efficacy in humans. The combination of various immunization routes and the use of suitable antigen-delivery systems may accomplish an important task-the induction of mucosal immune responses at a location relevant to the site of entry of a given pathogen.

Animals↗
PubMed1996

Systemic and mucosal protective immunity to pneumococcal surface protein A.

To date our studies demonstrate that PspA is a highly immunogenic molecule in mice and that it can elicit immunity to otherwise fatal infections following iv, ip, in, and it challenge. Although the molecule is serologically variable, it is sufficiently cross-reactive so that immunization with a single PspA can protect against strains of highly diverse serotypes. It is anticipated that a vaccine composed of a mixture of carefully chosen PspA molecules will be able to elicit protective immunity to virtually all pneumococci. If this vaccine proved efficacious in man, it would provide a more simple and less costly means of immunizing against pneumococcal infection than using recombinant vaccines. This could be especially important in the developing world where the cost of successful vaccines must be no more than pennies per dose. If PspA is found to be less efficacious than capsular polysaccharides, it may be valuable as a protein component of a PS-protein conjugate vaccine. In this capacity, PspA might expand the breath of protection elicited by a vaccine composed of only a few polysaccharide-protein conjugates representing capsule types most commonly associated with infectious pneumococci.

Animals↗
PubMed1996

A 500-kb physical map and contig from the Harvey ras-1 gene to the 11p telomere.

A contiguous physical map was constructed from the Harvey ras-1 (HRAS1) gene to the 11p telomere. The contig spans approximately 500 kb and is minimally composed of a telomere-containing YAC and P1 and cosmid clones. Included in the contig are 11 sequence-tagged sites derived from P1 and cosmid ends. Three genes were placed on the contig in the following order: telomere-ribonuclease/angiogenin inhibitor (RNH)-Harvey ras-1 (HRAS1)-HRAS1-related complex (HRC). Two novel tetranucleotide repeats (heterozygosity of 66 and 68%) and a complex CA repeat (heterozygosity of 78%) were isolated and characterized.

Base Sequence↗
PubMed1996

Chromosomal localization of 15 ion channel genes.

Several human Mendelian diseases, including the long-QT syndrome, malignant hyperthermia, and episodic ataxia/myokymia syndrome, have recently been demonstrated to be due to mutations in ion channel genes. Systematic mapping of ion channel genes may therefore reveal candidates for other heritable disorders. In this study, the GenBank and dbEST databases were used to identify members of several ion channel families (voltage-gated calcium and sodium, cardiac chloride, and all classes of potassium channels). Genes and ESTs without prior map localization were identified based on GDB and OWL database information and 15 genes and ESTs were selected for mapping. Of these 15, only the serotonin receptor 5HT3R had been previously mapped to a chromosome. A somatic cell hybrid panel (SCH) was screened with an STS from each gene and, if necessary the results verified by a second SCH panel. For three ESTs, rodent derived PCR products of the same size as the human STS precluded SCH mapping. For these three, human P1 clones were isolated and the genomic location was determined by metaphase FISH. These genes and ESTs can now be further evaluated as candidate genes for inherited cardiac, neuromuscular and psychiatric disorders mapped to these chromosomes. Furthermore, the ESTs developed in this study can be used to isolate genomic clones, enabling the determination of each transcript's genomic structure and physical map location. This approach may also be applicable to other gene families and may aid in the identification of candidate genes for groups of related heritable disorders.

Animals↗
PubMed1996

Construction and oral immunogenicity of a Salmonella typhimurium strain expressing a streptococcal adhesin linked to the A2/B subunits of cholera toxin.

A major adhesin from the oral pathogen Streptococcus mutans has been shown to be mucosally immunogenic upon genetic fusion with the cholera toxin A2/B subunits. To take advantage of the ability of Salmonella typhimurium to deliver cloned antigens to the mucosal inductive sites that would obviate the need for antigen purification, we expressed this chimeric construct in an attenuated S. typhimurium strain under the control of bacteriophage T7 transcription. Residual expression of the temperature-regulated T7 RNA polymerase at 30 degrees C allowed production of the chimeric protein at 2-3% of the total soluble protein, but it was increased five to six times following induction at 37 degrees C. Oral administration of a single dose of 10(9) recombinant Salmonella to mice resulted in serum IgG and salivary IgA antibody responses to Salmonella, cholera toxin, and the streptococcal adhesin, which were generally enhanced after a booster immunization.

Adhesins, Bacterial↗
PubMed1996

Induction of antibody-secreting cells and T-helper and memory cells in murine nasal lymphoid tissue.

Intranasal (i.n.) immunization is an effective route for inducing mucosal immune responses especially in the upper respiratory tract and mouth. To characterize the cells involved in these responses, nasal lymphoid tissue (NALT; considered to be the equivalent of Waldeyer's ring in humans) of normal mice, and of mice immunized intranasally with a bacterial protein antigen conjugated to cholera toxin B subunit, was isolated and the lymphoid cells analysed according to surface phenotype, immunoglobulin and antibody secretion, and cytokine profile. Compared with cells obtained from Peyer's patches (PP), NALT cells contained a higher proportion of T cells, especially naive (CD45RB+hi) T-helper cells, and fewer surface (s)IgA+ cells. Both tissues contained high proportions of sIgM+ IgD+ unswitched B cells. After i.n. immunization, IgA antibody-secreting cells were increased, indicating that isotype switching and differentiation of B cells to IgA-secreting cells occurred in NALT, whereas smaller numbers of antibody-secreting cells were found in PP after intragastric (i.g.) immunization. Antigen-specific memory cells persisted in NALT for at least 8 months after initial immunization. The cytokine expression profiles of antigen-stimulated NALT and PP cells of immunized mice, revealed by reverse transcription polymerase chain reaction analysis of mRNA, were similar. Both NALT and PP cells tended to express type 2 earlier or for longer than type 1 cytokine mRNA, but NALT cells tended to express interleukin-4 (IL-4) earlier, and IL-5 for a longer period, than PP cells. Thus NALT shares with PP cell populations typical of a mucosal inductive site, including unswitched B cells and naive T-helper (Th) cells. After i.n. immunization, NALT has the capacity to provide help for B-cell maturation and differentiation, as well as to maintain immune memory.

Animals↗
PubMed1996

KVLQT1 mutations in three families with familial or sporadic long QT syndrome.

Congenital long QT syndrome (LQTS) is a heterogeneous group of heritable disorders characterized by prolongation of the QT interval on the electrocardiogram, ventricular arrhythmias and sudden death. At least four genes can, when mutated, produce this phenotype. Of these genes, the recently identified KVLQT1 potassium channel is thought to be the one most commonly responsible. In this study, we used single strand conformational polymorphism (SSCP) analysis to screen two large and nine small LQTS families for mutations of the KVLQT1 potassium channel gene. We identified a novel missense mutation in two unrelated families which substitutes a serine for a conserved glycine in the putative pore region of the KVLQT1 channel. In a third family, a new alanine to valine mutation at a CpG dinucleotide resulted in the spontaneous occurrence of the long QT syndrome in monozygotic twin offspring of unaffected parents. Mutations at this same nucleotide have been observed in eight of the 19 LQTS families determined to have KVLQT1 mutations, suggesting this is a mutational hot spot. Both of these mutations alter the amino acid sequence in, or adjacent to, the pore of the channel and many diminish the channel's ability to conduct a repolarizing potassium current. To date, all KVLQT1 mutations determined to cause the LQTS are missense mutations. These data confirm the role of KVLQT1 in the LQTS and suggest that mutant KVLQT1 proteins may exert a dominant negative effect on repolarizing potassium currents by forming multimers with normal potassium channel protein subunits, dramatically reducing the number of fully-functional KVLQT1 channels.

Amino Acid Sequence↗