Darlene Charboneau

Immunogenicity and safety of pneumococcal vaccination in patients with rheumatoid arthritis or systemic lupus erythematosus.

Prevention of bacterial infection, which is a leading cause of morbidity in patients with rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE), is a priority. However, the safety and immunogenicity of the pneumococcal vaccine in such patients remain controversial. We evaluated the currently available pneumococcal vaccine in patients with RA or SLE. Pneumococcal vaccination was not associated with an appreciable deterioration in any clinical or laboratory measure of disease activity in either group. One month after vaccination, patients in both groups had significant increases in geometric mean concentrations of pneumococcal polysaccharide-specific IgG to all 7 serotypes tested, as did control subjects. However, 14 (33.3%) of 42 patients with RA and 5 (20.8%) of 24 patients with SLE responded either to none or to only 1 of the 7 polysaccharides. Pneumococcal vaccination is generally safe and immunogenic in patients with RA or SLE, but a subset of patients may remain unprotected by the currently available vaccine.

Magnitude, Duration, Quality, and Function of Pneumococcal Vaccine Responses in Elderly Adults

The suboptimal efficacy of the currently available 23-valent pneumococcal vaccine in the growing population of adults >65 years old may be related to the limited immunogenicity of the vaccine polysaccharides in this group. In this study, the majority of elderly outpatients with stable chronic illnesses generated a vigorous IgG response to seven vaccine serotypes comparable to that of healthy young adults at 1, 3, and 16 months after immunization. Moreover, the quality and function of anticapsular antibodies, measured as avidity and in vitro opsonization, were comparable between elderly and young subjects over time. However, a subset (approximately 20%) of elderly outpatients responded to fewer than two of seven serotypes tested 1 and 3 months after immunization, whereas none of the healthy young adults were such poor responders. Thus, despite the adequate mean immune responses of the elderly as a group, a substantial proportion of elderly persons may have poor responses to the currently available pneumococcal vaccine.

Dual function of pneumolysin in the early pathogenesis of murine pneumococcal pneumonia.

Streptococcus pneumoniae is one of the most common etiologic agents of community-acquired pneumonia, particularly bacteremic pneumonia. Pneumolysin, a multifunctional cytotoxin, is a putative virulence factor for S. pneumoniae; however, a direct role for pneumolysin in the early pathogenesis of pneumococcal pneumonia has not been confirmed in vivo. We compared the growth of a pneumolysin-deficient (PLY[-]) type 2 S. pneumoniae strain with its isogenic wild-type strain (PLY[+]) after direct endotracheal instillation of bacteria into murine lungs. Compared with PLY(-) bacteria, infection with PLY(+) bacteria produced greater injury to the alveolar-capillary barrier, as assayed by albumin concentrations in alveolar lavage, and substantially greater numbers of PLY(+) bacteria were recovered in alveolar lavages and lung homogenates at 3 and 6 h after infection. The presence of pneumolysin also contributed to the development of bacteremia, which was detected at 3 h after intratracheal instillation of PLY(+) bacteria. The direct effects of pneumolysin on lung injury and on the ability of pneumococci to evade local lung defenses was confirmed by addition of purified recombinant pneumolysin to inocula of PLY(-) pneumococci, which promoted growth of PLY(-) bacteria in the lung to levels comparable to those seen with the PLY(+) strain. We further demonstrated the contributions of both the cytolytic and the complement-activating properties of pneumolysin on enhanced bacterial growth in murine lungs using genetically modified pneumolysin congeners and genetically complement-deficient mice. Thus, pneumolysin facilitates intraalveolar replication of pneumococci, penetration of bacteria from alveoli into the interstitium of the lung, and dissemination of pneumococci into the bloodstream during experimental pneumonia. Moreover, both the cytotoxic and the complement-activating activities of pneumolysin may contribute independently to the acute pulmonary injury and the high rates of bacteremia which characterize pneumococcal pneumonia.

Pneumococcal IgA1 protease subverts specific protection by human IgA1

Bacterial immunoglobulin A1 (IgA1) proteases may sabotage the protective effects of IgA. In vitro, both exogenous and endogenously produced IgA1 protease inhibited phagocytic killing of Streptococcus pneumoniae by capsule-specific IgA1 human monoclonal antibodies (hMAbs) but not IgA2. These IgA1 proteases cleaved and reduced binding of the the effector Fcα1 heavy chain but not the antigen-binding F(ab)/light chain to pneumococcal surfaces. In vivo, IgA1 protease-resistant IgA2, but not IgA1 protease-sensitive IgA1, supported 60% survival in mice infected with wild-type S. pneumoniae. IgA1 hMAbs protected mice against IgA1 protease-deficient but not -producing pneumococci. Parallel mouse sera with human IgA2 showed more efficient complement-mediated reductions in pneumococci with neutrophils than did IgA1, particularly with protease-producing organisms. After natural human pneumococcal bacteremia, purified serum IgG inhibited IgA1 protease activity in 7 of 11 patients (64%). These observations provide the first evidence in vivo that IgA1 protease can circumvent killing of S. pneumoniae by human IgA. Acquisition of IgA1 protease-neutralizing IgG after infection directs attention to IgA1 protease both as a determinant of successful colonization and infection and as a potential vaccine candidate.

Studies of ammonia loading: effects of rate of delivery and enhanced removal of NH4 on blood levels of ammonia and coma induction.

Using dose-responce curves, the dose of NH4Ac inducing coma in one-half of the animals was increased by 60 to 80% after 1 mmol of arginine. The larger increase occurred in larger rats but was not proportional to the increase in weight. Incremental subcoma doses of NH4 raised the amount of NH4 required for inducing coma and the brain level of ammonia at the point of coma. After a portacaval shunt the results were similar, although lower doses of NH4 were required from the beginning. Blood ammonias after a loading dose (1.25 mmol) of NH4 were influenced by the duration of a preinfusion of NH4 and by the preinjection of various amino acids involved in the disposal of NH4 in the urea cycle. The amount of reduction in blood ammonia by ornithine and arginine compounds was less the longer the preinfusion of NH4. Blood ammonia was not lowered by glutamate at any time but was increased with longer preinfusion periods. Hepatectomy (Hx) reduced the removal of an NH4 load. After a modest load (0.85 mmol) of NH4, blood ammonia increased 5-fold, over that of shamoperated rats, with 70% Hx and 15-fold with 90% Hx. Ornithine reduced these blood ammonias by about 50%. Arginine had no effect. These studies indicate ways of reducing toxicity of NH4 and factors that predispose to or enhance toxicity.

4.5 Interaction of Bacteria and their Products with Tissue Culture Cells

Publisher Summary This chapter discusses interaction of bacteria and their products with tissue culture cells. Two general considerations apply to the use of cultured cells with bacteria— namely, avoiding contamination of cell cultures and using cells at a consistent phase of growth. Occult contamination of cell cultures can cause erratic cell growth and experimental results that cannot be repeated. Overt contamination of cell cultures causes loss of valuable time and occasionally loss of valuable cell lines. The second general consideration, culturing cells to a consistent phase of growth, requires that cells be split and seeded consistently for experiments. Typically, cells are used just before reaching confluence, equivalent to growth covering approximately 80% of the growing surface, to ensure that the cells are in log phase growth. Further, the chapter discusses the use of polarized, differentiated, and multiple layer cell culture systems. Multiple layer cell culture systems attempt to incorporate the complexities of interactions between barrier cells and supporting cells, such as epithelial and endothelial cell bilayer, or endothelial and smooth muscle cell bilayers.