Shirley J. Prince

Variations in immunoglobulins and IgA subclasses of human uterine cervical secretions around the time of ovulation.

The quantity and subclass distribution of IgA produced by the human uterine cervix may have a significant impact on the defence against sexually transmitted diseases as well as the regulation of fertility. Cervical mucus was obtained from 15 normal ovulating women around the time of ovulation. The total amounts of secreted IgA (including IgA1 and IgA2), IgG, and IgM were determined by ELISA. IgA was detected at high levels in all samples of cervical mucus. When ovulation was ascertained by daily urinary luteinizing hormone testing, IgA production was maximal 2–3 days before ovulation. Equal proportions of IgA1 and IgA2 were detected in cervical mucus, and 80% of the IgA occurred in the polymeric forms. The increased levels of IgA, the ratios of IgA1 to IgA2, and the predominance of polymeric IgA indicate that much of the IgA in human uterine cervical fluid originates from local production.

A Plastic Intraoral Device for the Collection of Human Parotid Saliva

A plastic intraoral (10) cup developed to facilitate collection of human parotid saliva was tested on two groups of patients. Samples collected in both IO and tubed cups exhibited no significant differences in lysozyme or lactoperoxidase activity, levels of total protein or secretory IgA, or in flow rates. Results suggest that the IO cup provides a simple, reliable method for collecting parotid saliva.

Intraperitoneal immunization of human subjects with tetanus toxoid induces specific antibody-secreting cells in the peritoneal cavity and in the circulation, but fails to elicit a secretory IgA response

Five patients on continuous ambulatory peritoneal dialysis (CAPD) were immunized intraperitoneally with tetanus toxoid (TT) through an indwelling catheter. Four control patients on CAPD received the same dose of TT intramuscularly. Before immunization, virtually no anti‐TT antibody‐secreting cells (AbSC) were detected by the enzyme‐linked immunospot (ELISPOT) assay in peripheral blood or peritoneal fluid from patients of either group. One to 2 weeks after immunization, high frequencies of TT‐specific AbSC were detected in the circulation and peritoneal cavity. More than 80% of those cells were of the IgG isotype, with IgA accounting for most of the remainder. Patients receiving TT by the i.p. route showed significantly higher frequencies of specific IgG and IgA AbSC in the peritoneal cavity than patients immunized intramuscularly. Frequencies of AbSC in peripheral blood did not significantly differ between the two groups. Immunization with TT by both routes resulted in a significant increase of IgG anti‐TT antibodies in serum, saliva and peritoneal fluid. A significant IgA antibody response was seen only in serum and peritoneal effluents. Therefore, i.p. immunization of human subjects with TT elicited both a localized response in the peritoneal cavity as well as a systemic response in serum, but did not induce a salivary IgA response.

Biosynthesis of J-chain in human lymphoid cells producing immunoglobulins of various isotypes

The relationship between synthesis, secretion, and subcellular localization of J-chain, IgM, IgA, and IgG was investigated in cultures of PWM-stimulated human PBL and in lymphoblastoid cell lines. Cells were examined for surface, cytoplasmic, and secreted immunoglobulins (Igs) and J-chain by immunofluorescence and radioimmunoassay (RIA). By these techniques, J-chain was detected in cells that produce polymeric or monomeric Igs. In PWM-stimulated PBL the synthesis of J-chain paralleled the production of Igs. In both PWM-stimulated (for 2 days) and unstimulated PBL, equal proportions of free and disulfide-linked J-chain were found. Increased amounts of intracellular J-chain were produced at later stages in PWM-stimulated PBL and J-chain occurred mostly in a free form. In tissue culture fluids, J-chain was not secreted in a free form but was always disulfide-linked to polymeric Igs. In lymphoblastoid cell lines, J-chain was present in a disulfide-linked form in IgM and IGA producers, but in IgG cells and in an IgM cell line (DAUDI) that did not secrete IgM but expressed it on the cell membrane, intracellular J-chain was present in free form. Although various proportions of polymeric and monomeric IgA were seen in culture fluids from IgA-secreting cell lines, intracellular IgA occurred mostly in a monomeric form. Further studies revealed that the ability to produce polymers was not equally distributed among all cells and might vary according to their content of J-chain and stage of maturation. Subcellular fractionation and subsequent analyses for J-chain and Ig in PWM-stimulated PBL and in IgM or IgG-producing cell lines revealed that these proteins were associated with fractions that contained ribosomes, cell sap, and low molecular weight RNA. In lysates of IgG and J-chain producing cells grown in the presence of 3H-labeled amino acids, intracellular J-chain was not disulfide-linked to IgG.

False positivity of enzyme-linked immunosorbent assay for measurement of secretory IgA antibodies directed at HIV type 1 antigens

We have determined that polymeric IgA in saliva of HIV-1-uninfected individuals binds in varying degrees to components of culture supernatants containing HIV-1 recombinant proteins when ELISA is used for the determination. This finding did not extend to salivary IgG antibodies. Further, such problems were not encountered in Western blot. Binding did not appear to be mediated by salivary proteins known to bind to IgA, including secretory component, amylase, lactoferrin, lysozyme, galactosyl transferase, or secretory leukocyte protease inhibitor, and was not influenced by blocking reagents or by changes in secondary anti-IgA antibodies. Although these findings will not likely impact on the use of saliva as a diagnostic fluid for HIV-1 infection (the HIV-1 response in saliva is mostly of the IgG isotype), they indicate that assessments of this secretion as an indicator of IgA mucosal immune responses to HIV-1 vaccines should be undertaken with caution.

IgA Antibodies to Oral and Ocular Bacteria in Human External Secretions

Abstract Human saliva contains secretory IgA (sIgA) antibodies to Streptococcus mutans ; however, lower antibody levels to serotype c than other serotypes of S. mutans are found. Oral challenge with S. mutans c and d resulted in implantation of c but not d , suggesting that naturally occurring salivary IgA antibodies regulate S. mutans colonization. Saliva, tears and milk contain naturally occurring sIgA antibodies to S. mutans and ocular bacteria, implying that these bacteria stimulate gut-associated lymphoreticular tissue (GALT) leading to sIgA responses in distant mucosal sites. Oral administration of S. mutans c to humans induced elevated levels of sIgA antibodies in external secretions and lowered numbers of S.mutans in dental plaque. Peripheral blood lymphocyte cultures stimulated with pokeweed mitogen and S. mutans gave antigen-specific IgA responses. These results show that ingested bacterial antigens stimulate GALT, and precursor-IgA B cells from this tissue migrate via the circulation to distant mucosal tissues; supporting the concept of a common mucosal IgA immune system in humans.

Molecular-cellular interactions in the secretory IgA system.

1) SC receptors were not detected on the surface of human PBL before or after PWM stimulation or on the surface of established lymphoblastoid cell lines. 2) SC binding was detected in the cytoplasm of differentiated lymphoid cells. The majority of the SC-binding cells contained intracellular IgA. 3) The binding of polymeric IgA to the surface of human epithelial cells (colonic carcinoma HT-29) was dependent on the presence of SC. 4) These findings indicate that SC is a receptor and possible transport protein for polymeric immunoglobulins, but that it is not directly involved in the homing of the IgA precursor cells to secretory tissues.

Intraperitoneal Administration of Tetanus Toxoid Elicits a Specific Response of Antibody-Secreting Cells in the Peritoneal Cavity

Anatomical studies suggested that the mammalian peritoneum plays an important role in immunological processes. Attention has primarily focussed on the greater omentum where “milky spots” cells play a role in the immunological defense of the peritoneal cavity.1,2,3,4,5 The peritoneal route of immunization has been used in experimental animals as an effective site for induction of both systemic and mucosal immune responses. 6,7,8 Kroese et al.9 showed that B cells from the murine peritoneal cavity repopulated the intestinal lamina propria of recipient mice with IgA-secreting cells. It was estimated that up to 50% of murine intestinal IgA-secreting cells were derived from surface IgA-negative precursor in the peritoneal cavity. Recently, Solvason et al.10 have demonstrated that human fetal omentum may serve as an additional site of B cell generation. These findings prompted us to study the potential of human peritoneal B cells to differentiate into antibody-secreting cells (AbSC). Patients on continuous ambulatory peritoneal dialysis (CAPD) represent a group of human subjects with a permanent access to the peritoneal cavity through an indwelling catheter.11 We immunized patients on CAPD intraperitoneally (i.p.) with tetanus toxid (TT), a well established protein antigen, to examine whether it can elicit specific antibody production by peritoneal B cells.

Serum and Salivary IgA1, IgA2, IgG and IgM Antibodies to Bacterial Antigens in Aged, Edentulous, and Normal Adults

Serum and saliva samples obtained from 51 aged edentulous (80 ± 4yr), 21 adult edentulous (47 ± 10yr), and 27 normal adult (39 ± 8yr) subjects, were assayed by ELISA for antibodies to Streptococcus mutans protein antigen I/II (Ag I/II), Escherichia coli K235 lipopolysaccharide (LPS), and phosphocholine-albumin (PC). Mean total salivary IgA levels were higher in the aged group, and some individuals had considerably elevated levels of IgA1. The levels of salivary IgA1 and IgA2, and serum IgA1, IgA2, IgG and IgM antibodies to Ag I/II and LPS were not signifcantly different between the aged and adult edentulous groups, nor between edentulous and normal adult groups. However, both edentulous groups had lower levels of serum IgM antibodies to PC than the normal subjects. Assuming that salivary antibodies reflect those found generally in the mucosal immune system, these results do not support the concept that mucosal IgA antibodies decline in the elderly

Immunological Properties and Differentiation Potential of Human Colostral Lymphocytes of B Cell Lineage

Human colostrum and milk contain approximately 1–5×106 cells/ml, identified by histochemical and immunohistochemical staining as macrophages (~40% of total cells), polymorphonuclear leukocytes (PMN), (~50%), lymphocytes (7–25%), and epithelial cells (~1%)1–4. Phenotypic analyses of lymphocytes revealed the presence of both B and T cells; the latter population is represented by CD4- and CD8-positive cells as well as γ/δ cells and NK cells2,5–7. Studies of colostral and milk lymphocytes of B cell lineage yielded controversial results with respect to their morphological properties, and their ability to respond in vitro to various stimuli and secrete immunoglobulins (Ig)1,2,4,8. The purpose of our studies was to re-evaluate the characteristics of colostral Ig-containing cells and to determine the transformability of B cells by the Epstein-Barr virus (EBV) which has been used in many studies of the differentiation potential of human B cells from various sources9.