Local production of secretory IgA in the eye-associated lymphoid tissue (EALT) of the normal human ocular surface.

PURPOSE: Secretory IgA (SIgA) is a critical local defense mechanism of mucosal immunity. Although the conjunctiva, as part of the ocular surface, has a mucosa-associated lymphoid tissue, the production of SIgA by local plasma cells and its transport is unequivocally accepted to occur only in the upstream lacrimal gland (LG). The molecular components were therefore investigated by immunohistochemistry (IHC) and their local production verified by RT-PCR. METHODS: Tissues from 18 conjunctivas and 9 LGs of human donor eyes with normal ocular surfaces were analyzed by histology and IHC. Different zones of 12 further conjunctivas and LG tissues were analyzed by RT-PCR for the presence of the respective mRNA. RESULTS: Plasma cells were present in the diffuse lymphoid tissue of all investigated specimens and showed an intense immunoreactivity for IgA. This immunoreactivity was absent when the antiserum was preadsorbed with the protein. The luminal epithelium, with the exception of goblet and basal cells, was strongly positive for the epithelial transporter molecule secretory component (SC) in the conjunctiva and interconnecting excretory duct similar to the LG. PCR products for IgA, the monomeric IgA-joining molecule (J-chain) and SC were regularly found in all conjunctival zones and in the LG in gel electrophoresis and were sequenced. CONCLUSIONS: The local production of SIgA is for the first time verified by RT-PCR in the human conjunctiva and in the LG. This finding points to an active role of the conjunctiva in secretory immune protection of the ocular surface and supports the presence and importance of EALT at the normal ocular surface.

Tumour necrosis factor-alpha-mediated human polymeric immunoglobulin receptor expression is regulated by both mitogen-activated protein kinase and phosphatidylinositol-3-kinase in HT-29 cell line.

Human polymeric immunoglobulin receptor (pIgR) is present on the surface of glandular epithelium, and it plays a crucial role in the mucosal immune defence. pIgR expression in HT-29 cells is up-regulated by one of the proinflammatory cytokines, tumour necrosis factor (TNF)-alpha. However, the mechanism used by the TNF-alpha-mediated signalling pathway has not been examined exclusively. To elucidate this mechanism in detail, HT-29 cells were cotreated with TNF-alpha and mitogen-activated protein kinase kinase (MAPKK, also called MEK1) inhibitor, PD98059, and the amount of free secretory component (SC) secreted into the culture medium was measured. The amount of free SC stimulated by TNF-alpha was increased by addition of PD98059. This up-regulation occurred at the transcriptional level. The amount of SC was also up-regulated by addition of TNF-alpha with U0126, an inhibitor of MEK1 and MEK2. Nuclear factor (NF)-kappaB activity and NF-kappaB binding to the kappaB2 site localized upstream of the pIgR gene did not change after coincubation of HT-29 cells with TNF-alpha and PD98059. The expression level of pIgR by TNF-alpha was decreased by LY294002, an inhibitor of phosphatidylinositol-3-kinase (PI3K), at the transcriptional level. Extracellular signal-regulated kinase (ERK)1/2 phosphorylation and NF-kappaB binding to the kappaB2 site were not affected by LY294002 treatment. These data suggest that TNF-alpha-mediated pIgR expression is negatively regulated by ERK pathway, which is independent of NF-kappaB. In addition, decrease of SC production by Ly294002 suggests that the presence of PI3K mediated regulation of SC production.

Preganglionic parasympathectomy decreases salivary SIgA secretion rates from the rat submandibular gland.

Immunoglobulin A (IgA) is transported into saliva by salivary cells expressing the polymeric immunoglobulin receptor (pIgR). In rat salivary glands, autonomic nerves stimulate this process. To examine how nerves affect pIgR-mediated IgA secretion, the chorda-lingual nerve was sectioned. One week after preganglionic parasympathectomy, both the stimulated and unstimulated rates of salivary IgA secretion were reduced, despite similar glandular amounts of IgA. Biochemical analysis of cells from parasympathectomised and control glands indicated reduced membrane expression of pIgR. It appears the removal of long-term parasympathetic input has affected the routing of pIgR within salivary cells and reduced the SIgA transport into saliva.

Lymphoid-epithelial secretory immune system in human fetuses in the second trimester of gestation.

The development of the secretory immune system (SIS) in the respiratory, digestive, and urogenital tracts and other organs of fetuses in the second trimester of gestation is described. Tissues of all internal organs of human fetuses (n = 36) that had died between 13 and 25 weeks of gestation were studied immunohistochemically for the presence of secretory component (SC), J chain, IgA, IgM, IgG, macrophages, and different subsets of lymphocytes. We found protein elements of the SIS in fetuses during the entire second trimester in the epithelium of the digestive, respiratory, and urinary tracts; in hepatocytes; in the epithelium of the bile duct, renal tubules, and all the urinary tract; in the salivary glands, pancreas, and thyroid; in the epithelium of the Fallopian tubes and uterus; in the epididymis and the rete testes; in the skin; and in other organs. Immunocompetent cells, including IgA- and IgM-secreting cells, were located in these organs under the epithelium and sometimes between epithelial cells. In fetuses with acute infection, the number of immunocompetent cells was higher, reflecting a whole-immune system reaction, including the SIS. We conclude that the fetal SIS is a ramified, defensive immune system that is distributed throughout most organs of epithelial origin in second-trimester fetuses, and that it reacts against intrafetal infiltration by foreign antigens.

Pneumococcal carriage and otitis media induce salivary antibodies to pneumococcal surface adhesin a, pneumolysin, and pneumococcal surface protein a in children.

Local antibodies probably contribute to defense against Streptococcus pneumoniae. This study examined whether pneumococcal carriage and acute otitis media (AOM) induce mucosal antibodies to potential vaccine candidates pneumococcal surface adhesin A (PsaA), pneumolysin (Ply), and pneumococcal surface protein A (PspA). IgA to all 3 proteins was detected by EIA in saliva of 329 children at ages 6, 12, 18, and 24 months and of 17 adults. A higher proportion of IgA-positive samples and higher antibody concentrations were seen in children with pneumococci-positive cultures of nasopharyngeal samples or middle ear fluid than in children with all cultures negative for pneumococci. The strong correlation between IgA and the presence of the secretory component suggests that the IgA was secretory. The findings indicate that pneumococcal carriage and AOM induce local production of anti-PsaA, anti-Ply, and anti-PspA antibodies early in life.

Reduced epithelial expression of secretory component in small airways correlates with airflow obstruction in chronic obstructive pulmonary disease.

The epithelial polymeric immunoglobulin receptor/transmembrane secretory component (pIgR/SC) transports into secretions polymeric immunoglobulin A (pIgA), which is considered the first line of defense of the respiratory tract. The present study, done with quantitative immunohistochemistry, evaluated epithelial expression of secretory component (SC) and Clara cell protein (CC16) and neutrophil infiltration into the airways of eight patients with severe chronic obstructive pulmonary disease (COPD) who were undergoing lung transplantation, as compared with these processes in six nonsmoking patients with pulmonary hypertension who were used as controls and in lung specimens from five smokers without chronic bronchitis. Staining for SC was significantly decreased in the COPD patients as compared with the controls, both in large (mean optical density [MOD]: 23.4 [range: 21.1 to 27.8] versus 42.2 [range: 28.2 to 49.3], p = 0.003) and in small airways (MOD: 30.8 [range: 20.3 to 39.4] versus 41.5 [range: 39.2 to 46.2], p = 0.003). SC expression in small airways correlated strongly with functional parameters such as FEV1 (Kendall's tau (K) = 0.76, p = 0.008), FVC (K = 0.64, p = 0.03), and midexpiratory flow at 50% of VC (MEF50) (K = 0.74, p = 0.01). The reduced expression of SC in large airways correlated with neutrophil infiltration in submucosal glands (K = -0.47, p = 0.03). Expression of CC16 in the bronchial epithelium of COPD patients was also significantly decreased as compared with that of controls, especially in small airways (MOD: 28.3 [range: 26.8 to 32.4] versus 45.8 [range: 40.7 to 56.0], p = 0.002), but no correlation was observed with lung function tests. In conclusion, this study shows that reduced expression of SC in airway epithelium is associated with airflow obstruction and neutrophil infiltration in severe COPD.

Role of local cytokines in increased gastric expression of the secretory component in Helicobacter pylori infection.

Using immunohistochemical staining, we examined the presence of secretory component (SC) on epithelial cells in gastric and duodenal biopsy specimens collected from Helicobacter pylori-infected individuals and healthy controls. Gastric epithelial cells from healthy volunteers expressed low, but detectable, levels of SC. In contrast, significantly higher level of expression of SC (P < 0.001) was observed on epithelial cells in the antra of H. pylori-infected individuals. The antral SC expression correlated with staining for gamma interferon of intraepithelial and lamina propria lymphocytes (r(s) = 0.76 and 0.69, respectively, P < 0.001) and correlated weakly with production of tumor necrosis factor alpha (r(s) = 0.43, P < 0.05), but it did not correlate at all with interleukin-4 production.

Different regulatory pathways employed in cytokine-enhanced expression of secretory component and epithelial HLA class I genes.

The transmembrane secretory component (SC, or pIg receptor) plays a crucial role in mucosal immunity by translocating dimeric IgA and pentameric IgM through exocrine epithelia. This receptor is up-regulated by cytokines in parallel with increased epithelial HLA expression. By use of the human epithelial cell line HT-29m3, we show that IFN-gamma, TNF-alpha and IL-4 activate transcription of the SC gene. This activation was slow, suggesting mediation via newly synthesized protein factors. IFN-gamma and TNF-alpha, but not IL-4, also up-regulated expression of HLA class I genes. However, this gene induction was rapid and did not depend on new protein synthesis. Nuclear run-on experiments showed that the transcription rate of HLA class I genes nearly peaked after only 30 min of IFN-gamma or TNF-alpha stimulation, whereas the SC transcription rate did not peak until after 20-36 h of IFN-gamma, TNF-alpha or IL-4 stimulation. Gel electrophoresis mobility shift assays demonstrated binding of nuclear proteins from cytokine-stimulated HT-29 cells to consensus elements in the promoter of the SC gene, involving the binding site for the nuclear factor-kappaB p50 subunit after TNF-alpha stimulation, and IFN-stimulated response element after IFN-gamma stimulation (and weakly after TNF-alpha. Our observations in vitro likely parallel events in vivo by which activated mucosal T cells and macrophages enhance pIg receptor-mediated external transport of secretory IgA and IgM and up-regulate epithelial HLA expression.

Production of secretory component and pathogenesis of gastric cancer in Helicobacter pylori-infected stomach.

The purpose of this study was to examine the production of secretory component (SC) and immunoglobulin A (IgA) in the gastric mucosa with Helicobacter pylori infection and to investigate the influence of immunological reactions on various phases of infection (gastritis, intestinal metaplasia, gastric cancer). Production of SC and IgA was assessed by immunohistochemical staining in (1) endoscopic biopsy samples of H. pylori-eradicated cases (n = 25), and (2) surgically resected stomach tissues of H. pylori-positive gastric cancer cases, intestinal type (IGC, n = 25) and diffuse type (DGC, n = 25). Before eradication therapy, all samples showed positive staining of SC and IgA in epithelial cells, and IgA was also positive in plasma cells in the mucosal layer. H. pylori bacteria were positively stained for SC and IgA. After treatment, the degree of SC and IgA staining in epithelial cells was reduced with successful eradication; but with intestinal metaplasia, SC staining was positive regardless of the results of treatment. In nonmetaplastic mucosa, SC-positive cells were increased in the glandular neck zone to the surface mucosal layer; and the intensity of SC staining was increased in proportion to the degree of mucosal inflammation and IgA-positive cell aggregation. In intestinal metaplasia, SC was positive irrespective of the degree of inflammation. Most cancer foci also showed positive staining of SC, irrespective of histological type. Production of SC and IgA was thought to be a specific reaction against H. pylori infection, occurring from the early to the late stages and not limited to intestinal metaplasia. It was suggested that immunological reactions against H. pylori infection might generally be involved with the pathogenesis of intestinal metaplasia and both histological types of gastric cancer (IGC and DGC).

Secretory component production by human bronchial epithelial cells is upregulated by interferon gamma.

Secretory immunoglobulin A (S-IgA) participates in the first noninflammatory line of defence of the respiratory tract. S-IgA consists of dimeric IgA (dIgA) produced by plasma cells and secretory component (SC) produced by epithelial cells. This study compared SC production by primary cultures of human bronchial epithelial cells (HBEC) and by respiratory epithelial cell lines. Among the cell lines, A549 did not produce detectable SC, 16HBE produced very low levels of SC, while CALU-3 produced significant levels of SC. HBEC produced SC in nonpolarized and polarized primary cultures, where it was secreted apically. Polarized HBEC transcytosed radiolabelled and cold dIgA, resulting in the presence of S-IgA in their apical media. SC production and IgA transcytosis by polarized HBEC were upregulated by interferon-gamma (IFN-gamma) after 48 h. By reverse transcription-polymerase chain reaction, no SC messenger ribonucleic acid (mRNA) was detected in A549 and 16HBE, while SC mRNA in CALU-3 was comparable to that of HBEC incubated for 48 h with IFN-gamma. By immunocytochemistry, HBEC expressed SC immunostaining and its intensity increased after 48 h with IFN-gamma. It is concluded that human bronchial epithelial cells produce secretory component and transcytose dimeric immunoglobulin A in vitro. These processes were apically polarized and upregulated by interferon-gamma. Among the cell lines studied, only CALU-3 expressed secretory component-messenger ribonucleic acid and produced detectable secretory component.

Uterine stromal cell suppression of pIgR production by uterine epithelial cells in vitro: a mechanism for regulation of pIgR production.

Previous studies have shown that the polymeric Ig receptor (pIgR) is produced by rat uterine epithelial cells both in vivo and vitro. The expression of the pIgR is regulated by sex hormones and/or cytokines at mucosal sites, however the mechanism of regulation in the uterus is not clear. In these studies, co-culture of stromal cells from mature rat uteri with uterine epithelial cells decreased epithelial cell pIgR production. Conditioned supernatants from stromal cells incubated with epithelial cells also decreased pIgR production. Immunohistochemical studies confirmed that expression of pIgR on uterine epithelial cells decreased in the presence of stromal cells. Viability of epithelial cells was sustained during these experiments, as evidenced by the maintenance of high transepithelial resistance. These studies are the first report of stromal cell regulation of pIgR production by epithelial cells at any site in the body and suggest that stromal cells can provide a signal that leads to the regulation of pIgR production.

Production of secretory immunoglobulin A by a single mammalian cell.

Secretory IgA (sIgA) plays a critical role in providing protection against infection at the mucosal surfaces. Normally, sIgA is the product of two different cell types with heavy, light, and J chains produced by the plasma cells, whereas secretory component (SC), a cleavage product of the polymeric immunoglobulin receptor (pIgR), is added during the transit of dimeric IgA through the epithelial cell layer. In the current study, by introducing a gene for the processed form of SC into a cell line that produces dimeric IgA, we have succeeded in creating a single cell that is able to produce and secrete covalently joined sIgA. To our knowledge, this is the first time it has been possible to efficiently produce large quantities of sIgA of defined specificity in mammalian cells. The sIgA made using this approach has great potential as an immunotherapeutic.

The mechanism of Epstein-Barr virus infection in nasopharyngeal carcinoma cells.

To investigate the relationship between Epstein-Barr virus (EBV) and nasopharyngeal carcinoma (NPC) cells, we examined the pathway of EBV infection in NPC cell lines. We used immunolocalization to investigate the EBV receptor (C3d-R) and polymeric immunoglobulin receptor [secretory component (SC) protein]. We incubated IgA anti-EBV and EBV particles with NPC cells and observed the EBV DNA signal by in situ polymerase chain reaction hybridization and polymerase chain reaction plus Southern blotting. We also colocalized SC protein and EBV RNA in NPC biopsy specimens. Results showed that: 1) NPC cells did not express the EBV receptor but did express SC protein in each line; 2) SC protein was also expressed in some tumor cells but not in untransformed squamous metaplastic epithelia in NPC biopsy specimens; 3) EBV could infect NPC cells through an EBV-IgA and SC complex and retained an EBV viral genome in their nuclei; SC expression could be down-regulated by EBV proteins; and 4) in biopsy specimens, a fraction of tumor cells showed SC protein expression; only a portion of tumor cells contained EBV, and of these cells only a few expressed SC protein. These findings indicate that EBV cannot infect untransformed nasopharyngeal squamous metaplastic epithelia but can enter NPC cells through IgA-mediated endocytosis.

Effects of biliary obstruction on hepatic deoxyribonucleic acid and protein synthesis after partial hepatectomy.

BACKGROUND/AIMS: Major hepatectomy for biliary malignancy associated with biliary obstruction is often complicated due to hepatic failure. To determine the effects of biliary obstruction on liver regeneration, hepatic deoxyribonucleic acid and protein synthesis after partial hepatectomy was studied in rats with pre-operative biliary obstruction. MATERIALS AND METHODS: After 5 or 14 days of biliary obstruction, experimental rats underwent concurrent biliary decompression and 70% partial hepatectomy. Control rats underwent partial hepatectomy alone. Hepatic deoxyribonucleic acid synthesis was determined 24 hours after partial hepatectomy. Hepatocellular protein synthesis and secretory protein synthesis were determined at baseline and 48 hours after partial hepatectomy. RESULTS: Deoxyribonucleic acid synthesis was significantly depressed in rats with biliary obstruction compared with controls (p < 0.05). Baseline hepatocellular protein synthesis and secretory protein synthesis, especially secretory protein synthesis, increased in proportion to the duration of biliary obstruction (p < 0.01). After partial hepatectomy, control rats showed marked increases in hepatocellular protein synthesis and secretory protein synthesis (p < 0.01). Increased baseline hepatocellular protein synthesis and secretory protein synthesis were preserved in the regenerating liver of rats with biliary obstruction. CONCLUSIONS: The results suggest that biliary obstruction gives priority to secretory protein synthesis over DNA synthesis which may inhibit liver regeneration.

Effect of nicotine on secretory component synthesis by secretory epithelial cells.

Previously, we reported that secretory component (SC), lactoferrin (LF), and lysozyme (LY) levels were significantly lower in saliva from smokeless tobacco (ST) users than in saliva from control non-tobacco users. However, the levels of salivary immunoglobulin A were significantly higher, albeit with an altered attachment of SC, in ST users than in control subjects. SC, LF, and LY are synthesized by secretory epithelial cells at mucosal sites adjacent to lymphocyte regions. In the present report, HT-29 human epithelial cells, cultured with various concentrations of an ST aqueous extract or pure nicotine (0 to 1 mg/ml) or cotinine (0 to 5 mg/ml), exhibited significantly lower levels of cell-associated cell lysate (CL) and secreted culture supernatant (CS) SC, LF, and LY than cells cultured without ST components. Nicotine significantly decreased (P < or = 0.05) the synthesis of SC by 20 to 100%, LF by 20 to 60%, and LY by 5 to 75% of CL and CS control values. Studies also indicated significant decreases (P < or = 0.05) in SC, LF, and LY levels in both CL and CS of cells cultured with ST aqueous extract or cotinine. Total cell numbers and metabolic activity significantly decreased primarily when cells were incubated with higher concentrations of ST extract, nicotine, or cotinine. The addition of human recombinant interleukin-4 or gamma interferon diminished the effects ST had on HT-29 cell synthesis of SC, LF, and LY. Our data indicate that nicotine, cotinine, and ST have an adverse effect on synthesis and secretion of SC, LF, and LY. These effects were below ST concentrations found to be cytotoxic for secretory epithelial cells. Furthermore, addition of interleukin-4 or gamma interferon reduced the suppressive effect of ST on synthesis or secretion of SC, LF, or LY.

Secretory component mRNA and protein expression in colorectal adenomas and carcinomas.

Secretary component (SC) is expressed basolaterally as a transmembrane protein (pIg receptor) on secretory epithelial cells. As pIg receptor it plays a central role in humoral immunity by mediating the external translocation of dimeric IgA and pentameric IgM. A few case reports have suggested that reduced or absent SC protein expression is associated with diarrhoeal disease, but there is no convincing evidence that a primary pIg receptor deficiency can occur. In this study the relative presence of SC mRNA was determined by Northern blot analysis and related to immunohistochemically determined SC protein expression in 33 colorectal adenomas (31 patients) with increased risk of developing sporadic colorectal cancer, as well as in 19 colorectal carcinomas from 19 patients with such sporadic tumours. In the adenomas, SC mRNA levels were positively related to SC protein expression; both mRNA and SC protein were negatively related to histological grade. Similarly, SC mRNA levels tended to be related to the SC protein expression in the carcinomas. SC mRNA was detected in all adenomas, and only two of ten carcinomas (10.5%) deemed to be SC deficient by immunohistochemistry also lacked SC mRNA expression, suggesting diallelic alterations in the SC-encoding gene (locus PIGR). This possibility agreed with Southern blot analysis performed on a separate sample of 32 other colonic carcinomas in which the diallelic loss of D1S58 (which exhibits a close linkage centromerically to PIGR) was calculated to be 6.4%. Together these findings suggested that reduced SC protein expression in colorectal adenomas might be a transcriptional defect reflecting the degree of cellular dysplasia, whereas absent SC protein expression in colorectal carcinomas might also involve post-transcriptional defects and occasional diallelic gene deletions representing late events in carcinogenesis.

Secretory component (polymeric immunoglobulin receptor) expression on human keratinocytes by stimulation with interferon-gamma and differences in response.

Secretory IgA (sIgA) is a major protective factor in the mucosal immune system because of its great ability to form complexes with bacteria. Secretory component (SC) is an 80-kDa glycoprotein, a component of sIgA, which functions as a polymeric immunoglobulin receptor for IgA and aids the secretion of sIgA from the epithelial surface. We studied SC production by keratinocytes which were involved in the inflammatory process using interferon-gamma (IFN-gamma) as one of the major inflammatory promoters produced by helper T cells. Using two human squamous cell carcinoma cell lines (HSCs) and normal human keratinocytes (NHKs), results from flow cytometric analysis, enzyme-linked immunosorbent assay (ELISA), and Northern blotting revealed that HSCs produced SC when stimulated with IFN-gamma, although their responses differed; one line exhibited enhanced SC production whereas the production in the other line was suppressed. NHKs also exhibited SC expression on the cell surface by means of immunocytochemical analysis, flow cytometry and ELISA, however the responses were also different in each strain. Although the reason for the diversity of SC expression on keratinocytes is not clear, these differences may influence epidermal sIgA secretion level.

Lymphocyte adhesive interactions with lacrimal gland acinar epithelial cells in primary culture.

PURPOSE: In lacrimal glands, cell-cell interactions control the localization of lymphocyte populations that play a role in immune defense at the ocular surface. This study describes lymphocyte adhesive interactions with cultured lacrimal gland acinar epithelial cells. METHODS: Primary cultures of lacrimal gland epithelial cells were used as targets for in vitro lymphocyte binding assays. The relative adherence of lymphocyte populations was determined. Various physiologically active agents and putative ligand analogs were tested for their effect in the binding assay. RESULTS: Thoracic duct lymphocytes (TDL) bound to cultured lacrimal acinar epithelial cells in greater numbers than did thymocytes (54 cells/mm2 versus 8 cells/mm2). B cells showed preferential adherence compared with T cells (75% sIg+, 14% W3/13+). Thoracic duct lymphocyte binding required intact metabolic and membrane-cytoskeletal function and was inhibited by treating the lymphocytes with sodium azide, formaldehyde, or cytochalasin B (23%, 12%, and 10% of control binding, respectively). Further, adherence was dependent on divalent cations. Ethylenediaminetetraacetic acid-mediated inhibition (42% of untreated) was restored by replacing calcium (89%) but not magnesium (41%). Lymphocyte adherence was inhibited in the presence of fucoidin or phosphomannan polysaccharides (36% and 48% of control binding, respectively). Fibronectin peptides, which are involved in certain types of integrin-mediated adherence, had no effect in this system. Lacrimal culture supernatants contained a factor that was inhibitory for TDL adherence (more than 50% inhibition when concentrated 5 or 10 times). CONCLUSIONS: Thoracic duct lymphocyte adherence to cultured lacrimal gland acinar epithelial cells shows good correlation with previously reported adherence to lacrimal gland frozen sections. Further, lacrimal cell culture supernatants contain soluble factors that inhibit TDL adherence to epithelial cells. These findings suggest that the lacrimal molecules involved in lymphocyte localization are shed and that lacrimal epithelial cell cultures will be useful for ligand isolation and characterization.

Production of human secretory component with dimeric IgA binding capacity using viral expression systems.

The cDNA encoding the NH2-terminal 589 amino acids of the extracellular domain of the human polymeric immunoglobulin receptor was inserted into transfer vectors to generate recombinant baculo- and vaccinia viruses. Following infection of insect and mammalian cells, respectively, the resulting truncated protein corresponding to human secretory component (hSC) was secreted with high efficiency into serum-free culture medium. The Sf9 insect cell/baculovirus system yielded as much as 50 mg of hSC/liter of culture, while the mammalian cells/vaccinia virus system produced up to 10 mg of protein/liter. The M(r) of recombinant hSC varied depending on the cell line in which it was expressed (70,000 in Sf9 cells and 85-95,000 in CV-1, TK- 143B and HeLa). These variations in M(r) resulted from different glycosylation patterns, as evidenced by endoglycosidase digestion. Efficient single-step purification of the recombinant protein was achieved either by concanavalin A affinity chromatography or by Ni(2+)-chelate affinity chromatography, when a 6xHis tag was engineered to the carboxyl terminus of hSC. Recombinant hSC retained the capacity to specifically reassociate with dimeric IgA purified from hybridoma cells.