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At least 19 recordsLinked to original sources
PubMed1989

Quantification of human urinary free secretory component, secretory and nonsecretory IgA by ELISA.

The specific quantification of human urinary free secretory component (FSC), secretory IgA (SIgA) and total IgA using ELISA has been hampered by mutual interferences of these three molecules. Using affinity chromatographically purified antisera an attempt was therefore made to reduce these interferences without necessitating further assay steps. FSC and total IgA were measured in unprocessed urine by means of anti-FSC and anti-IgA as well as alkaline phosphatase-coupled anti-FSC or anti-IgA antisera. SIgA was determined using anti-IgA as well as alkaline phosphatase-coupled anti-FSC. Nonsecretory urinary IgA was calculated from the measured SIgA and total IgA. The mutual interferences of FSC, SIgA or nonsecretory IgA in the three assay systems were low and not relevant for normal samples. Normal urinary concentrations were: FSC 344 +/- (SD) 208 ng/ml (n = 120), SIgA 1,874 +/- 1,133 ng/ml (n = 123) and nonsecretory IgA, depending on the way of standardization, 712 +/- 699 (n = 56) or 878 +/- 732 ng/ml (n = 51). SIgA excretion increased with age. Lower urinary SIgA as well as total and nonsecretory IgA levels were observed in males as compared to females. No correlation evolved between the hormonal status of women and the excretion of FSC, SIgA or IgA. In IgA-deficient patients virtually no nonsecretory IgA or SIgA was detected in the urine while the FSC concentration was in the normal range.

Adult↗
PubMed1984

Characterization of epitopes of human secretory component on free secretory component, secretory IgA, and membrane-associated secretory component.

We have compared the epitopes present in various forms of human secretory component by using a panel of hybridoma-derived antibodies elicited by immunizing mice with free secretory component (FSC) or secretory IgA (sIgA). Enzyme-linked immunosorbent binding assays (ELISA) were used to assess antibody binding to FSC- and SC-containing antigens, including sIgA isolated from milk, reduced and alkylated sIgA, and sIgA assembled in vitro by incubating dimeric IgA with FSC. Immunofluorescence assays were also used to assess binding to a human epithelial tumor cell line (HT29) that expresses secretory component as an integral protein of the plasma membrane. The results can be summarized as follows. 1) Most antibodies from fusions in which sIgA was the immunizing antigen bound preferentially to sIgA. 2) Most antibodies from fusions in which FSC was the immunizing antigen bound preferentially to FSC. 3) Antibodies that bound preferentially to sIgA invariably bound sIgA assembled in vitro; antibodies that bound preferentially to FSC invariably did not. 4) Antibodies that bound readily to both sIgA and FSC were rare in all fusions. 5) The monoclonal antibodies defined at least six classes of epitopes on SC, including epitopes that were a) FSC specific and reduction sensitive, b) FSC specific and reduction insensitive, c) sIgA specific and reduction-sensitive, d) sIgA specific and reduction insensitive, e) shared by FSC and sIgA and reduction-sensitive, and f) shared by FSC and sIgA and reduction-insensitive. 6) Antibodies that mediated intense immunofluorescent staining of secretory component on HT29 cell membranes were rare and constituted a distinct subset of those which recognized epitopes shared by FSC, reduced and alkylated sIgA, and some preparations of native sIgA. Results of these antibody-binding studies indicate that most SC epitopes are not shared by FSC and sIgA. Most SC-related epitopes on sIgA appear to be generated by the physical interaction of SC with dimeric IgA, whereas most epitopes on FSC are masked or altered by this interaction. Finally, epitopes that are shared by membrane SC and FSC and/or sIgA represent a minor and immunochemically distinct subset of epitopes on SC. The high proportion of unique epitopes on the different physical forms of SC suggest that the epitopes of this molecule are highly sensitive to its molecular environment. The monoclonal reagents described here will be useful in studying the structure and function of SC; quantitating FSC, sIgA, and membrane SC; purifying various molecular forms of SC by immunoaffinity chromatography; and localizing SC in human tissues and cultured cells by immunocytochemical techniques.

Alkylation↗
PubMed1991

Characterization of a critical binding site for human polymeric Ig on secretory component.

Secretory component (SC) is an integral membrane glycoprotein of secretory epithelial cells which is responsible for the specific transport of polymeric Ig (PIg) to external mucosal surfaces. The ectoplasmic segment which binds polymeric Ig is comprised of five Ig-type domains. Chemically and enzymatically modified forms of the ectoplasmic portion of SC (FSC) were produced and tested for their ability to bind to PIgA and PIgM. Deglycosylated FSC bound specifically to PIg, indicating that N-linked carbohydrate moieties on FSC are not required for binding. Denatured, reduced, and alkylated FSC did not bind to PIgA, and bound to PIgM with significantly reduced affinity, suggesting that native conformation of the polypeptide backbone of SC was important to binding. Tryptic fragments of FSC which bound to PIg were isolated and identified to be derived from domain I of SC. Synthetic peptides comprising overlapping portions of domain I bound to PIg to varying degrees. The strongest affinity was demonstrated by a peptide comprised of residues 15 to 37 of SC. A comparison of the amino acid sequences of human, rabbit, and rat SC indicated that this region contained a high degree of residue identity (78%) and may represent a consensus sequence for binding of FSC to PIg. Importantly, the peptide comprised of residues 15 to 37 was also recognized by a monoclonal antibody, 6G11, which inhibited the binding of FSC to PIgA. These results demonstrate that the binding of human SC to PIg is critically dependent on a highly conserved peptide region within the first domain of SC centering at residues 15-37.

Amino Acid Sequence↗
PubMed1991

The usefulness of routine screening for salivary secretory component.

Secretory IgA is a dimeric immunoglobulin found in association with the J chain and secretory component (SC). It is secreted into saliva and other mucosal fluids and is involved in mucosal immunity. The absence of either SC or secretory IgA may be associated with recurrent sinopulmonary infections, diarrhea, and failure to thrive. We present a retrospective study of 1262 samples from 877 patients who were screened for salivary IgA, SC, and serum immunoglobulin levels. Forty-six patients (5.2%) of those tested were found to have absent salivary SC. Although only 19 of these patients (41.3%) could be retested, all were found to have SC on repeated testing. Of the patients whose initial samples of saliva exhibited no SC, 15% (6/46) had low or absent serum IGA (less than 10 mg/dl) in contrast to 8.6% (66/769) of patients whose saliva contained detectable SC, but this was not statistically significant (chi 2 = 1.93; p greater than 0.1). There was also no correlation between serum immunoglobulin levels and the absence of SC. Because of the rarity of salivary SC deficiency, routine screening is not valuable.

Aging↗
PubMed1980

A radioimmunoassay for quantitation of secretory component.

Secretory component (SC) isolated from human colostrum was radioiodinated with chloramin-T method and it was subjected to a radioimmunoassay for quantitation of SC. A monospecific rabbit anti-sera against human SC which could react with free SC as well as immunoglobulin bound SC was used for the assay. The present method can measure as low as 300 ng/ml of SC. The level (mean +/- 2 S.D.) of serum SC in healthy subjects was determined to be 31.3 +/- 7.2 micrograms/ml. Studies with Sephadex G-200 gel filtration demonstrated that SC in the normal sera existed mostly as a form of SC-IgA. In contrast, free SC was never detected in them.

Colostrum↗
PubMed1987

Cleavage of membrane secretory component to soluble secretory component occurs on the cell surface of rat hepatocyte monolayers.

Rat liver secretory component is synthesized as an integral membrane protein (mSC) and cleaved to an 80-kD soluble form (fSC) sometime during transcellular transport from the sinusoidal to the bile canalicular plasma membrane domain of hepatocytes. We have used 24-h monolayer cultures of rat hepatocytes to characterize the conversion of mSC to fSC. Cleavage of mSC in cultured hepatocytes is inhibited by the thiol protease inhibitors leupeptin, antipain, and E-64, but not by other inhibitors, including disopropylfluorophosphate, pepstatin, N-ethylmalemide, p-chloromercuribenzoic acid, and chloroquine. Leupeptin-mediated inhibition of cleavage is concentration dependent and reversible. In the presence or absence of leupeptin, only 10-20% of mSC is accessible at the cell surface. To characterize the behavior of surface as opposed to intracellular mSC, cell surface mSC was labeled with 125I by lactoperoxidase-catalyzed iodination at 4 degrees C. Cell surface 125I-mSC was converted to extracellular fSC at 4 degrees C in the absence of detectable internalization. Cleavage was inhibited by leupeptin and by anti-secretory component antiserum. Cleavage also occurred at 4 degrees C after cell disruption. In contrast, 125I-mSC that had been internalized from the cell surface was not converted to fSC at 4 degrees C in either intact or disrupted cells. Hepatocytes metabolically labeled with [35S]cys also released small quantities of fSC into the medium at 4 degrees C. The properties of fSC production indicate that cleavage occurs on the surface of cultured rat hepatocytes and not intracellularly. Other features of the cleavage reaction suggest that the mSC-cleaving protease is segregated from the majority of cell surface mSC, possibly within a specialized plasma membrane domain.

Animals↗
PubMed1982

Characterization of epithelial elements in human major salivary glands by functional markers: localization of amylase, lactoferrin, lysozyme, secretory component, and secretory immunoglobulins by paired immunofluorescence staining.

Human parotid and submandibular glands were studied by paired immunofluorescence staining, including a variety of combinations of fluorochrome conjugates with contrasting colors. Lactoferrin (Lf), secretory component (SC), and particularly amylase were demonstrated in serous acinar cells of both glands. In addition, lysozyme (Ly) was present in some acini, although mainly located in intercalated ducts where Lf was also most commonly seen. SC was present in acini, intercalated ducts, and striated ducts but not in large collecting ducts. Staining for SC was generally faint but increased in intensity at the cell periphery and particularly at the luminal face of striated duct cells. Immunoglobulin (Ig) A--and IgM when detectable--showed an epithelial distribution similar to that of SC, in accordance with the known secretory properties of these two Ig classes. Conversely, IgG was not present in epithelial cells, despite its high extravascular concentrations. Mucous epithelial elements did not show unequivocal staining for any of the proteins studied. Formaldehyde fixation, combined with pronase treatment of tissue sections and prolonged exposure (20 hr) to antibody, enhanced markedly the staining intensity for lysozyme; ethanol fixation and 30-min incubation with conjugates generally afforded better localization of the other epithelial components.

Adult↗
PubMed1984

Rat liver membrane secretory component is larger than free secretory component in bile: evidence for proteolytic conversion of membrane form to free form.

Secretory component is a receptor for polymeric immunoglobulins on epithelial cells and hepatocytes that facilitates transport of polymeric immunoglobulins into external secretions. Little is known about the transcellular migration of secretory component-polymeric IgA complexes or the membrane forms of secretory component. We therefore examined rat bile and liver membranes to identify and compare the various molecular species of secretory component. Bile or liver membrane proteins were electrophoresed in sodium dodecyl sulfate-polyacrylamide gels and electrophoretically transferred to nitrocellulose membranes. Protein profiles on blots were probed with antisecretory component antiserum, and the immunoreactive bands were visualized by indirect immunoperoxidase staining. Bile collected in the presence of proteolytic inhibitors showed an immunoreactive doublet band (Mr = 82,000 and 78,000) in the molecular weight range of free secretory component. By contrast, free secretory component in bile collected in the absence of proteolytic inhibitors and purified by affinity chromatography migrated as a single protein with an Mr = 70,000. Both components of the free secretory component doublet bound dimeric IgA when blots were probed with human dimeric IgA. Crude liver membranes prepared in the presence of proteolytic inhibitors showed two immunoreactive secretory component-containing bands, Mr = 107,000 and 99,000, whereas membranes prepared without proteolytic inhibitors showed two smaller immunoreactive bands; one of these proteolytically severed proteins comigrated with the 82,000-dalton free secretory component in bile. These results indicate that membrane forms of secretory component are present in rat liver. The observations that the membrane secretory component is larger than biliary free secretory component and yields biliary SC-like forms of secretory component upon proteolysis support the hypothesis that free secretory component in bile is a proteolytic product of larger liver membrane-associated secretory component.

Animals↗
PubMed1998

Simplified method for purification of colostrum to obtain secretory component of immunoglobulin A, using secretory component as a reference protein in tracheal aspirate fluid.

Many studies employ bronchoalveolar lavage fluid for assessment of biologically active substances secreted from the lung. However, investigators continue to search for a useful reference standard to correct for the inevitable but variable degree of dilution of this fluid. The glycoprotein, soluble secretory component of IgA, may serve as a valid reference protein. We report a simplified method for the purification of secretory component from colostrum. Soluble secretory component was isolated from human colostrum using serial centrifugation, size-exclusion fractionation and ion-exchange chromatography. Secretory component rich fractions were assayed by enzyme immunoassay. They were also evaluated for total amino acid content and distribution and sequence determination with satisfactory agreement with published results. We then demonstrated that soluble secretory component concentration in tracheal aspirate fluid did not correlate with either albumin or with total protein measured in the same samples. Therefore, we conclude that the secretory component of IgA serves as a useful reference marker because its use may avoid errors resulting from leakage of plasma proteins into epithelial lining fluid. Advantages of this method for establishing a standard for secretory component include ready availability of soluble secretory component, simplicity of the method and relative rapidity of the techniques.

Chemical Fractionation↗
PubMed1987

Purification of polymeric immunoglobulin from cell culture supernatants by affinity chromatography using secretory component.

Human secretory component bound covalently to Sepharose 4B has been used as an affinity adsorbent to isolate and purify polymeric immunoglobulin from cell culture supernatants. The method was used to isolate murine IgM isotype anti-trinitrophenol antibody and rat IgM isotype anti-lymphocyte antibody from hybridoma cell culture supernatants. Gel filtration of the eluted antibodies followed by enzyme immunoassay showed that all recovered IgM was of pentameric molecular size. Murine IgA isotype anti-dinitrophenol antibody and murine IgA anti-human rotavirus antibody were isolated from cell culture supernatants of a plasmacytoma and a hybridoma respectively. Most of the IgA recovered following affinity adsorption with secretory component was of greater molecular size than dimer. Murine IgG was not adsorbed by secretory component bound to Sepharose. Eluted antibody retained antigen binding activity. Affinity chromatography using human secretory component bound covalently to a solid phase provides an antigen-independent technique for purification of murine and rat IgA and IgM polymeric immunoglobulin from cell cultures.

Animals↗
PubMed1981

The membrane receptor for polymeric immunoglobulin is structurally related to secretory component. Isolation and characterization of membrane secretory component from rabbit liver and mammary gland.

The membrane receptor of epithelial cells which binds polymeric immunoglobulins and mediates their transepithelial translocation is antigenically related to secretory component (SC), a glycoprotein produced by the epithelial cells. In rabbit milk, secreted SC is found either bound to polymeric immunoglobulin A (IgA) or in a free form. It is comprised of a heterogeneous population of molecules with apparent Mr = 83,000 and 80,000 (upper doublet) and 58,000 and 55,000 (lower doublet). Membrane SC was isolated by immunoadsorption from deoxycholate-solubilized plasma membranes of rabbit liver and mammary gland. The purified proteins, heterogenous in size with apparent Mr = 120,000 and 116,000 (upper doublet) and 95,000 and 91,000 (lower doublet), were amphiphilic. One-dimensional peptide maps revealed extensive structural homology between both the upper doublets of the membrane and secreted SC, as well as between the lower doublets. This indicates that the high Mr membrane SC of hepatocytes and mammary cells are precursors of the lower Mr secreted SC found in bile and milk. All four membrane SC, purified in high yield from rabbit liver by immunoadsorption and preparative electrophoresis in the presence of sodium dodecyl sulfate, bound IgA dimer specifically. The affinity (Kd approximately equal to 10 nM) was similar to the one observed in free SC-IgA dimer interaction. Only the membrane proteins recognized by the anti-SC antibodies were able to bind polymeric IgA. In liver membrane preparations, the receptor was localized at the surface of smooth vesicles, using IgA dimer-biotin and avidin-gold. Our results indicate that the high Mr membrane SC is the plasma membrane receptor for polymeric immunoglobulin.

Animals↗
PubMed1995

Human eosinophils express a receptor for secretory component. Role in secretory IgA-dependent activation.

The existence of a functional receptor for secretory component (SC) on the eosinophil membrane might explain the preferential degranulation induced by secretory IgA (sIgA) when compared to serum IgA. Indeed, flow cytometry analysis revealed that purified human SC could bind to a subpopulation (4-59%) of blood eosinophils purified from 19 patients with eosinophilia. Binding of radiolabeled human SC could be competitively inhibited using unlabeled SC or secretory IgA but not with serum IgA or IgG. Immunoprecipitation and immunosorbent chromatography using human SC revealed the presence of a major component at 15 kDa in eosinophil extracts as well as in culture supernatants but not in neutrophils. The 15-kDa protein eluted from the human SC immunosorbent was able to bind to SC or to sIgA but not to serum IgA. Eosinophils preincubated with human SC or sIgA released eosinophil cationic protein (ECP) and eosinophil peroxidase (EPO) after addition of anti-SC or anti-IgA monoclonal antibody as respective cross-linking reagents. These results indicated that binding of free or complexed SC to human eosinophils could induce eosinophil degranulation. Furthermore, the dose-dependent inhibition by SC of mediator release induced by sIgA but not by serum IgA, suggested that the receptor for SC could be involved in the preferential degranulation mediated by sIgA. These results indicate a novel pathway of eosinophil activation and its potential involvement in mucosal immunity, particularly in inflammatory diseases associated with infiltration of eosinophils and the enhanced production of sIgA.

Binding, Competitive↗
PubMed1980

Studies on secretory component in digestive disease. III. Levels of serum secretory component in digestive disease.

The levels of serum secretory component (SC) were measured in 147 patients with digestive disease. Decreased levels were found patients with acute hepatitis, HBs-antigen associated chronic hepatitis, HBs-antigen associated liver cirrhosis and hepatoma. Normal levels were observed in patients with diabetes mellitus, gastric cancer and colonic carcinoma. Elevated levels were found in patients with cholecystitis, obstructive jaundice and acute pancreatitis. The serum SC level in almost all disease groups showed no correlation with immunoglobulin levels.

Diabetes Mellitus↗
PubMed1980

Studies on secretory component in digestive disease. IV. Levels of serum secretory component in liver disease.

The levels of serum secretory component (SC), immunoglobulins, rheumatoid factor, HBs-antigen and HBs-antibody were determined in patients with liver disease, especially chronic liver disease. In chronic liver disease, decreases in serum SC were accompanied by increases in the titer of rheumatoid factor. Patients with chronic liver disease and HBs-antigen or HBs-antibody had decreased levels of serum SC compared to seronegative patients. There was no correlation between the levels of serum SC and immunoglobulin in patients with chronic liver disease.

Adult↗
PubMed1996

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.

Cell Line↗