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Biomedical subjects

M E Lamm

Publications and source records attributed to M E Lamm.

At least 19 recordsLinked to original sources
PubMed1996

Hepatic pathology resulting from mouse hepatitis virus S infection in severe combined immunodeficiency mice.

Mouse hepatitis virus (MHV) is a pervasive pathogen that causes morbidity and mortality in mouse colonies worldwide. Although it is not a major cause of mortality in immunocompetent mice, infections from MHV strains of lower virulence can be fatal to athymic nude mice. The histopathologic features and alterations of serum biochemical parameters resulting from infection with a low-virulence MHV strain in severe combined immunodeficiency (scid) mice has not been well described. Thus we recently studied the disease caused by MHV-S in scid mice after intranasal inoculation. Mouse hepatitis virus infection in scid mice, which have severe defects of B and T cells, may be highly lethal, resulting in immediate mortality. However, our results indicate that scid mice survived for an average of 12 to 14 days after infection with doses of MHV up to 10(7) PFU/mouse. The virus caused a significant increase in serum enzyme activities and bilirubin concentration associated with histologically demonstrable hepatocellular injury at postinoculation days 3, 4, and 8. Furthermore, virus was detected in mouse liver homogenates and nasal and bronchial lavage specimens. These results provide valuable information regarding the histopathologic and biochemical consequences of MHV-S infection in scid mice.

Alanine Transaminase↗
PubMed1995

Comparative studies of transcytosis and assembly of secretory IgA in Madin-Darby canine kidney cells expressing human polymeric Ig receptor.

Epithelial transport of polymeric IgA (pIgA) from its site of synthesis to the mucosal lumen is mediated by the polymeric Ig receptor (pIgR). During transcytosis, a disulfide bond forms between pIgR and pIgA, resulting in secretion of a covalently linked complex. To dissect further the intracellular processing and functions of pIgR, we have expressed the entire coding sequence of human pIgR cDNA in Madin-Darby canine kidney (MDCK) cells. Cloned transfected cells express human pIgR, as detected by immunofluorescence and by quantification of the cleaved extracellular domain of pIgR in culture supernatants. The function of transfected pIgR was confirmed by measuring vectorial transcytosis of 125I-labeled pIgA and its disulfide bonding to pIgR. Species specificity of transcytosis was determined by comparing transport of human, rat, and mouse pIgA in MDCK cells expressing either human or rabbit pIgR. pIgA from all three species was transported by both human and rabbit pIgR, with rat pIgA being transported to the greatest extent in each case. However, disulfide bonding was observed only with human pIgR, and was found to occur mainly inside the cell. Our results suggest that conformational differences between human and rabbit pIgR may account for differences in disulfide bonding to pIgA, and show that efficient transcytosis of pIgA is correlated better with noncovalent than covalent binding to pIgR.

Animals↗
PubMed1995

IgA and mucosal defense.

The traditional role of IgA antibodies in mucosal defense has been considered as providing an immune barrier to keep exogenous substances, including microbial pathogens, from penetrating the mucosa. In this way infections can be prevented. More recently, studies in vitro and in vivo are providing evidence to suggest that IgA may have additional roles in mucosal defense. For example, during their passage through the lining epithelial cells of mucous membranes en route to the secretions, IgA antibodies may have an opportunity to neutralize intracellular pathogens like viruses. Also, IgA antibodies in the mucosal lamina propria have opportunities to complex with antigens and excrete them through the adjacent mucosal epithelium, again by the same route to the secretions that is taken by free IgA. These latter functions could aid in recovery from infection.

Animals↗
PubMed1994

Epithelial transcytosis of monomeric IgA and IgG cross-linked through antigen to polymeric IgA. A role for monomeric antibodies in the mucosal immune system.

We recently demonstrated that the polymeric IgR (pIgR) mediates epithelial transcytosis of immune complexes (IC) containing dimeric IgA (dIgA). In vivo, this "excretory" pathway could allow direct elimination of IgA IC at the mucosal sites where they are likely to form. In contrast, IC containing only monomeric IgA (mIgA) were not transported, consistent with the specificity of pIgR for polymeric IgA. However, the potential exists in vivo that monomeric Ig like mIgA or IgG could become associated through binding to multivalent Ag with IC containing dIgA, and that such mixed IC could act as ligands for pIgR. In the present work, using Madin-Darby canine kidney epithelial cells that express pIgR, we showed that 125I-labeled anti-DNP mIgA or IgG in the same IC with unlabeled dIgA antibody and DNP-BSA Ag was vectorially transported from the basolateral to the apical surface and then released. However, 125I-mIgA IC or 125I-IgG IC (without dIgA) and 125I-mIgA or 125I-IgG in the presence of dIgA antibody (but without Ag) were not transported, demonstrating the necessity for monomeric Ig to be in an IC with dIgA to be transported. Transcytosis of mixed mIgA/dIgA or IgG/dIgA IC was mediated by pIgR because no transport was observed in untransfected, wild-type Madin-Darby canine kidney cells lacking pIgR. The data demonstrate that mIgA and IgG can participate along with dIgA in the "excretory" pathway for local elimination of IgA IC, thus providing a means by which monomeric antibodies have the potential to participate in the mucosal immune system.

Animals↗
PubMed1994

Disulfide bond formation between dimeric immunoglobulin A and the polymeric immunoglobulin receptor during hepatic transcytosis.

The polymeric immunoglobulin receptor on rat hepatocytes binds dimeric IgA on the sinusoidal surface and mediates its transport to the canaliculus, where the complex of dimeric IgA and secretory component, the cleaved extracellular domain of polymeric immunoglobulin receptor, is secreted into bile. This process is unique in that disulfide bonds are formed between dimeric IgA and polymeric immunoglobulin receptor during transcytosis, permanently preventing their dissociation. Here we present three lines of evidence that disulfide bonding between dimeric IgA and polymeric immunoglobulin receptor occurs predominantly in a late transcytotic compartment and that hepatic transcytosis can proceed in the absence of disulfide bond formation. First, throughout the course of transcytosis the percentage of intracellular dimeric IgA disulfide bonded to polymeric immunoglobulin receptor is less than half that in bile, suggesting that disulfide bond formation is a late event in transcytosis. Second, dimeric IgA that recycles from early endocytotic compartments into the circulation is mostly noncovalently bound to secretory component. Finally, the rate of transcytosis of dimeric IgA and its appearance in bile are not affected when disulfide bond formation with polymeric immunoglobulin receptor is inhibited by blocking of free thiol groups on dimeric IgA with iodoacetamide. These results are consistent with other findings in the literature and indicate that the main physiological role of disulfide bond formation between dimeric IgA and polymeric immunoglobulin receptor is not to facilitate transcytosis but, rather, to stabilize the dimeric IgA-secretory component complex after its release into external secretions such as bile and intestinal secretions.

Animals↗
PubMed1994

Experimental IgA nephropathy secondary to hepatocellular injury induced by dietary deficiencies and heavy alcohol intake.

BACKGROUND: In humans, alcoholic liver disease is frequently associated with IgA mesangial deposits, microscopic hematuria and a small amount of proteinuria, identifying a secondary form of IgA nephropathy. Alcoholic liver disease is almost always associated with nutritional deficiencies. EXPERIMENTAL DESIGN: In order to examine the relationship between alcohol intake and/or inadequate diet and IgA nephropathy, groups of 4 week-old-male Lewis rats were maintained on a lipotrope-deficient (LD) diet (N = 20), intragastric infusions of a commercial whiskey (1.5 ml/100 gm body weight) three times a week, and regular chow (N = 23) or both intragastric whiskey infusion and an LD diet (N = 17). A fourth control group (N = 19) was given no whiskey and normal chow. RESULTS: All rats given the LD diet had marked steatosis and elevated "liver" enzymes. Changes were more severe, and with early bridging fibrosis and nodule formation in those also given whiskey, associated with increased hepatic content of mRNA encoding transforming growth factor-beta. A moderate steatosis without alteration in serum enzymes or transforming growth factor-beta expression was found in rats given whiskey (all p < 0.0001) compared with controls. IgA accumulated in hepatic sinusoids instead of in canaliculi and bile ducts, suggesting impaired transport of IgA and IgA immune complexes from blood to bile, in rats given an LD diet and/or whiskey infusion. A moderate increase in mesangial matrix was observed only in rats given both whiskey and an LD diet. Bright granular IgA and mild granular C3 mesangial deposits and electron-dense deposits were evident in 63 to 70% of experimental rats (all p < 0.001) versus only trace deposits in 5 to 11% of controls. Moderate IgG codeposits were present in 34 to 55% of rats given the LD diet and/or whiskey (all p < 0.02), versus trace deposits in 10% of controls. Significant hematuria and proteinuria were observed in rats given the LD diet and/or whiskey (p < 0.0001) versus controls. Intestinal permeability measured by xylose absorption was significantly increased relative to controls only in rats given both whiskey and the LD diet (p < 0.001). Serum IgA specific for selected alimentary antigens was increased relative to controls in 75 to 100% of the experimental rats. CONCLUSIONS: The combination of LD diet and alcohol intake, which mimics the human alcoholic condition, promotes hepatic and renal changes, leading to hepatocellular injury and a secondary form of IgA nephropathy.

Alcoholism↗
PubMed1993

Unstable inter-H chain disulfide bonding and non-covalently associated J chain in rat dimeric IgA.

Disulfide bonds are a major force in stabilizing the three-dimensional structure of immunoglobulins. To determine the pattern of interchain disulfide bonding between the four H chains, four L chains and single J chain of rat dimeric IgA (dIgA), we analyzed dIgA from the LO DNP-64 hybridoma by diagonal SDS-PAGE. Bands corresponding to one, two, three and four H chains, one and two L chains and the free J chain were observed under non-reducing conditions, suggesting that the interchain disulfide bonds in rat dIgA are unstable under denaturing conditions. Similar patterns of disulfide bonding were observed in three other hybridoma or myeloma dIgAs from LOU/CN rats. In contrast, when dIgA pretreated with iodoacetamide (IA) was analyzed by the same technique, only bands corresponding to four H chains, one and two L chains and the free J chain were observed, suggesting that blocking free sulfhydryl groups stabilizes the inter-H chain disulfide bonds. Reaction of dimeric LO DNP-64 dIgA with 5,5'-dithiobis-(2-nitrobenzoic acid) or with 14C-IA demonstrated that this dIgA contains an average of 4 moles of free sulfhydryl groups per mole of protein under non-denaturing conditions and 9 moles of free sulfhydryl groups under denaturing conditions. Taken together, the results suggest that interchain disulfide bonds in rat dIgA are unstable, presumably due to the influence of nearby free sulfhydryl groups, and that non-covalent forces are critical for stabilizing the dIgA complex. The results also indicate that J chain is entirely non-covalently associated with the H chains, an apparently unique feature of rat dIgA. A model for interchain disulfide bonding in rat dIgA is proposed.

Animals↗
PubMed1993

A three-tiered view of the role of IgA in mucosal defense.

Mucosal IgA has generally been viewed as an immune barrier to prevent the adherence and absorption of antigens. Recent studies employing polarized epithelial monolayers have suggested two additional functions for mucosal IgA. One is to neutralize intracellular microbial pathogens, such as viruses, directly within epithelial cells. The second is to bind antigens in the mucosal lamina propria and excrete them through the adjacent epithelium into the lumen, thereby ridding the body of locally formed immune complexes and decreasing their access to the systemic circulation.

Animals↗
PubMed1992

Intracellular neutralization of virus by immunoglobulin A antibodies.

IgA is thought to neutralize viruses at the epithelial surface of mucous membranes by preventing their attachment. Since IgA, a polymeric immunoglobulin, is transported through the lining of epithelial cells by the polymeric-immunoglobulin receptor and since viruses are obligate intracellular parasites, we hypothesized that IgA antibodies may also interfere with viral replication by binding to newly synthesized viral proteins within infected cells. Polarized monolayers of Madin-Darby canine kidney epithelial cells expressing the polymeric-immunoglobulin receptor were infected on the apical surface with Sendai virus. Anti-Sendai virus IgA monoclonal antibody delivered from the basolateral surface colocalized with viral protein within the cell, as documented by immunofluorescence. More importantly, anti-viral IgA reduced virus titers greater than 1000-fold (P less than 0.0001) in apical supernatants and greater than 10-fold (P less than 0.0001) in cell lysates from monolayers treated with anti-viral IgA compared with those treated with either anti-viral IgG or an irrelevant IgA monoclonal antibody. We believe that the differences in viral titers between cell layers treated with specific IgA, which enters the epithelial cell by binding to the polymeric-immunoglobulin receptor, and those treated with specific IgG, which does not enter the cells, or irrelevant IgA indicate that specific intracellular IgA antibodies can inhibit viral replication. Thus, in addition to the classical role of humoral antibodies in extracellular defense, IgA antibody may be able to neutralize microbial pathogens intracellularly, giving IgA a role in host defense that has traditionally been reserved for cell-mediated immunity.

Animals↗
PubMed1992

Comparison of IgA versus IgG monoclonal antibodies for passive immunization of the murine respiratory tract.

The protective efficacy of anti-Sendai virus IgA was compared to that of IgG after topical application of monoclonal antibodies (MAb) to the respiratory tract of mice. BALB/c mice were passively intranasally immunized with 50 microliters ascites containing equivalent ELISA titers of MAb 1 h before and 4 and 24 h after intranasal challenge with Sendai virus. Lung viral titers were determined by plaque assay 3 days following challenge. In most instances IgA MAb afforded equivalent protection to IgG MAb in that there was no significant difference in virus recovery from the lungs of animals treated with either IgA or IgG MAb, including subclasses of IgG. When IgA MAb was fractionated into monomers and oligomers, there was no inherent advantage to the oligomeric form with respect to passive protection against viral challenge. The data indicate that IgA and IgG antibodies are equally efficacious in protecting the airways from viral infection. The experiments suggest that the advantage of IgA for protecting mucosal surfaces, such as the respiratory tract, relates to the presence of a specialized mechanism for transporting oligomeric IgA across epithelial surfaces. The results also support the rationale for active mucosal immunization protocols designed to generate an IgA response.

Animals↗
PubMed1992

Glomerular hemodynamics and eicosanoid synthesis in a rat model of IgA nephropathy.

We sought to study glomerular pathophysiology in a model of IgA nephropathy (IgAN). Preliminary experiments with oral immunization indicated that Lewis rats had higher IgA levels and IgA/IgG ratios than Wistar, Fischer or Sprague-Dawley rats. Six groups of Lewis rats were studied: four continuously orally immunized for eight weeks with bovine gamma globulin (BGG) in the drinking water, the other two non-immunized controls. Groups of immunized rats were treated with a thromboxane receptor antagonist (SQ 29,485) and/or a thromboxane synthase inhibitor (UK 38,485). After systemic challenge, microscopic hematuria was present in 84% of immunized rats not given anti-thromboxane drugs versus 18% of non-immunized rats (P less than 0.01). Immunized rats showed predominantly IgA glomerular deposits with lesser IgG and C3, and produced more glomerular thromboxane than controls, but no significant increase in prostaglandin E2. Immunized rats also had reduced GFR and RPF, but not a reduced filtration fraction, compared to controls. Thromboxane synthase inhibitor diminished glomerular thromboxane and increased prostaglandin E2 in immunized rats. Anti-thromboxane therapy reduced hematuria and apparently re-established the RPF but not the GFR in immunized rats, yielding a reduced filtration fraction. We propose that increased thromboxane, in concert with mesangial contraction that is unaffected by anti-thromboxane drugs, contributes to the pathophysiology in this model of IgAN.

Animals↗
PubMed1991

The polymeric immunoglobulin receptor (secretory component) mediates transport of immune complexes across epithelial cells: a local defense function for IgA.

The polymeric immunoglobulin receptor (pIgR) on mucosal epithelial cells binds dimeric IgA (dIgA) on the basolateral surface and mediates transport of dIgA to the apical surface. Using Madin-Darby canine kidney epithelial cells stably transfected with pIgR cDNA, we found that soluble immune complexes (ICs) of 125I-labeled rat monoclonal antidinitrophenyl (DNP) dIgA (125I-dIgA) and DNP/biotin-bovine serum albumin were transported from the basolateral to the apical surface and then released. Monomeric IgA ICs were not transported, consistent with the specificity of pIgR for polymeric immunoglobulins. Essentially all the 125I-dIgA in apical culture supernatants was streptavidin precipitable, indicating that dIgA remained bound to antigen during transcytosis. While both dIgA and dIgA ICs bound pIgR with equal affinity (Kd approximately 8 nM), the number of high-affinity binding sites per cell was 2- to 3-fold greater for dIgA than for dIgA ICs. The extent of endocytosis of dIgA and dIgA ICs was correlated with the number of high-affinity binding sites. SDS/PAGE analysis of intracellular dIgA and dIgA ICs demonstrated that in both cases IgA remained undegraded during transport. The results suggest that the pathways of epithelial transcytosis of free dIgA and dIgA ICs are the same. Given the high population density of mucosal IgA plasma cells and the enormous surface area of pIgR-expressing mucosal epithelium, it is likely that significant local transcytosis of IgA ICs occurs in vivo. Such a process would allow direct elimination of IgA ICs at the mucosal sites where they are likely to form, thus providing an important defense function for IgA.

Animals↗
PubMed1991

Cell polarity regulates the release of secretory component, the epithelial receptor for polymeric immunoglobulins, from the surface of HT-29 colon carcinoma cells.

The HT-29 human colon carcinoma cell line differentiates in glucose-free medium to an enterocytic phenotype. We previously isolated a series of HT-29 subclones selected for high levels of expression of secretory component (SC), the epithelial receptor for polymeric immunoglobulins. To develop a model system for studying effects of cell polarity on SC expression and release from the cell surface, the HT-29.74 subclone was induced to differentiate in glucose-free medium. Expression of SC was induced by glucose deprivation in both the parental HT-29 cell line and, to an even greater extent, in the HT-29.74 subclone. Prolonged glucose deprivation of HT-29.74 cells resulted in morphological changes consistent with enterocytic differentiation. Metabolic radiolabeling of SC in differentiated HT-29.74 cells indicated that proteolytic cleavage of membrane-bound to free SC occurred both on the cell surface and intracellularly, possibly in a vacuolar apical compartment or intrapeithelial lumen. To study effects of cell polarity on SC release, differentiated HT-29.74 cells were depolarized by culturing in low calcium medium. Within 2 hours after transfer of the cells into low calcium medium, a burst of SC release was observed concomitant with cell depolarization. Subsequently, release of SC declined significantly and remained low as long as cells were maintained in a depolarized state. The extent of cell depolarization could be controlled by varying the extracellular calcium concentration or by substituting the divalent cation Sr++, which partially prevents depolarization, for Ca++. In either case, the magnitude of the initial burst and subsequent decline in release of SC was proportional to the extent of cell depolarization. We conclude that cell polarity plays an important role in controlling the release of SC in intestinal epithelial cells, most likely by regulating the distribution of membrane-bound SC and SC protease, which are on the basolateral and apical cell surfaces, respectively, in differentiated cells.

Calcium↗