IgA Molecules Research Articles

IgA-specific proteins of pathogenic bacteria

Data on structure and specificity of bacterial IgA receptors (IgA-binding M-like proteins Arp4 and Sir22 from hemolytic streptococci of serogroup A, beta-antigen from hemolytic streptococci of serogroup B, and SSL family proteins from Staphylococcus aureus) are surveyed in this review. The principal conclusion derived from comparison is the fact that all bacterial receptors bind the same site in the IgA molecule overlapping with the binding site of endogenous human IgA receptor CD89. We assume that this site, consisting of spatially close amino acid strands Leu257-Gly259 in domain Calpha2 and Pro440-Phe443 in domain Calpha3, is subject to conformational rearrangement induced by the binding of antigen in the IgA active site.

Pathogenetic significance of aberrant glycosylation of IgA1 in IgA nephropathy

IgA nephropathy (IgAN), the most common form of primary glomerulonephritis worldwide, is defined by predominant IgA1 deposits in the glomerular mesangium. Among abnormalities of the IgA immune system reported so far in IgAN, aberrant O-linked glycosylation in the hinge region of IgA1 is the most consistent finding. IgA1 molecules bearing abnormal glycosylation have been found in serum, in tonsillar lymphocytes, and in eluate from mesangial deposits, and characterized by decreased O-linked N-acetylgalactosamine residues with or without alteration in the terminal sialylation of the O-linked sugars. IgA1 with incomplete galactosylation has a tendency to accumulate in glomerular mesangium by self-aggregation or immune complex formation. Glomerular mesangial cells exposed to immune complexes of these IgA1 can proliferate and secrete cytokines, chemokines, growth factors, and extracellular matrix components promoting inflammatory reactions in the glomeruli. Although genes encoding enzymes involved in the O-glycosylation process, such as C1GALT1, have been reported to be responsible for susceptibility to IgAN, recent evidence suggests that the abnormality is restricted to a small fraction of B cell populations and arises from dysregulated IgA1 production and secretion in mucosal immune system. This review will focus on and discuss the role of incompleteness of IgA1 O-galactosylation in the pathogenesis of IgAN and propose a possible mechanism in which abnormal IgA1 occurs in IgAN.

Recombinant human immunoglobulin (Ig)A1 and IgA2 anti‐D used for detection of IgA deficiency and anti‐IgA

To avoid anaphylactic reactions, immunoglobulin (Ig)A-deficient patients with anti-IgA should be transfused with IgA-deficient blood components. There is a need for fast and robust assays for demonstration of IgA deficiency and for detection of anti-IgA. Recombinant human IgA1 and IgA2 anti-D molecules were constructed, expressed in Chinese hamster ovary cells, and purified. These antibodies were used to sensitize group O D+ red blood cells (RBCs) for use as indicator cells, either in the format of a passive hemagglutination inhibition assay for detection of IgA deficiency or in a passive hemagglutination assay for detection of anti-IgA. Both assays were performed in gel card. The sensitivity for IgA detection was adjusted to approximately 100 ng per mL. The assay for demonstration of IgA deficiency correlated with an enzyme-linked immunosorbent assay for quantification of IgA. Anti-IgA were easily detected, and the reactivity with IgA anti-D-sensitized RBCs could be inhibited by purified IgA1 and/or IgA2 and by normal plasma containing IgA but not by IgA-deficient plasma. Anti-IgA was found in 64 percent of IgA-deficient donors with less than 3 ng of IgA per mL. The assays for detection of IgA and anti-IgA described in this article are fast and robust. Furthermore, they are applicable in all standard blood typing laboratories and are therefore well suited for immediate investigation of transfusion reactions.

Galactosylation of N- and O-linked carbohydrate moieties of IgA1 and IgG in IgA nephropathy.

The mechanism of IgA deposition in the kidneys in IgA nephropathy is unknown. Mesangial IgA is of the IgA1 subclass, and since no consistent antigenic target for the IgA1 has been described, we have investigated the glycosylation of the molecule, as a potential non-immunological abnormality which may contribute to its deposition. IgA1 is rich in carbohydrate, carrying N-linked moieties in common with IgG, but also O-linked sugars, which are rare in serum proteins, and not expressed by IgG or IgA2. Lectin binding assays were designed to examine the expression of terminal galactose on the N-linked carbohydrate chains of purified serum IgG and IgA1, and the O-linked sugars of IgA1 and C1 inhibitor (one of the very few other serum proteins with O-linked glycosylation). No evidence was found for abnormalities of N-linked glycosylation of either isotype in IgA nephropathy compared with matched controls. However, in IgA nephropathy, reduced terminal galactosylation of the hinge region O-linked moieties was demonstrated; this was not seen in C1 inhibitor, which showed normal or increased galactosylation of the O-linked sugars. This abnormality of IgA1 has considerable implications for the pathogenesis of IgA nephropathy, since the O-linked sugars lie in an important functional location within the IgA1 molecule, close to the ligand of Fc receptors. Changes in the carbohydrates in this site may therefore affect interactions with receptors and extracellular proteins, leading to anomalous handling of the IgA1 protein in this condition, including failure of normal clearance mechanisms, and mesangial deposition.

Nonhuman Primate IgA: Genetic Heterogeneity and Interactions with CD89

Nonhuman primates are extremely valuable animal models for a variety of human diseases. However, it is now becoming evident that these models, although widely used, are still uncharacterized. The major role that nonhuman primate species play in AIDS research as well as in the testing of Ab-based therapeutics requires the full characterization of structure and function of their Ab molecules. IgA is the Ab class mostly involved in protection at mucosal surfaces. By binding to its specific Fc receptor CD89, IgA plays additional and poorly understood roles in immunity. Therefore, Ig heavy alpha (IGHA) constant (C) genes were cloned and sequenced in four different species (rhesus macaques, pig-tailed macaques, baboons, and sooty mangabeys). Sequence analysis confirmed the high degree of intraspecies polymorphism present in nonhuman primates. Individual animals were either homozygous or heterozygous for IGHA genes. Highly variable hinge regions were shared by animals of different geographic origins and were present in different combinations in heterozygous animals. Therefore, it appears that although highly heterogeneous, hinge sequences are present only in limited numbers in various nonhuman primate populations. A macaque recombinant IgA molecule was generated and used to assess its interaction with a recombinant macaque CD89. Macaque CD89 was able to bind its native ligand as well as human IgA1 and IgA2. Presence of Ag enhanced macaque IgA binding and blocking of macaque CD89 N-glycosylation reduced CD89 expression. Together, our results suggest that, despite the presence of IgA polymorphism, nonhuman primates appear suitable for studies that involve the IgA/CD89 system.

Aberrantly Glycosylated IgA1 in Glomerular Immune Deposits of IgA Nephropathy

In IgA nephropathy, abnormal O-glycosylation of IgA1 molecules contributes to mesangial IgA1 deposition and the development of glomerular injury; however, direct in situ demonstration of aberrantly O-glycosylated IgA1 within glomerular immune deposits has not been reported. This study investigated the presence of abnormally glycosylated IgA1 in situ and its spatial relationship with complement within the immune deposits and correlated these features with glomerular lesion severity. Immunofluorescence and confocal microscopy were used to evaluate 19 consecutive renal biopsies, and the severity of glomerular lesions were also scored. Aberrantly glycosylated IgA was observed within the immune deposits, and its amount was found to correlate with both the severity of glomerular lesions and the amount of C3c on the surface of the deposits. These results demonstrate that qualitative and quantitative evaluation of aberrantly glycosylated IgA can be performed on routine renal biopsy samples. Its presence in immune deposits likely influences the spatial organization of IgA and C3c, thereby contributing to the glomerular inflammatory response in IgA nephropathy.

Immunopathogenesis of IgAN

The defining hallmark of IgA nephropathy (IgAN) is deposition of polymeric IgA1 in the glomerular mesangium accompanied by a mesangial proliferative glomerulonephritis. The mechanisms involved in mesangial polymeric IgA1 deposition and the initiation of inflammatory glomerular injury remain unclear. This lack of a complete understanding of the pathogenesis of IgAN has meant that there is still no treatment known to modify mesangial deposition of IgA. Increasing evidence, however, supports the importance of IgA-containing immune complex formation as a pivotal factor driving mesangial IgA deposition and triggering of glomerular injury. A number of potentially important changes to the IgA1 molecule have been identified in IgAN, which may contribute to immune complex formation. These changes suggest that the polymeric IgA1 that deposits in IgA nephropathy is derived from mucosally primed plasma cells. The presence of this IgA in the circulation reflects displacement of mucosal B lineage cells to systemic sites and may be the result of mishoming of lymphocytes trafficking along the mucosa-bone marrow axis.

Analysis of O-glycan heterogeneity in IgA1 myeloma proteins by Fourier transform ion cyclotron resonance mass spectrometry: implications for IgA nephropathy

IgA nephropathy (IgAN) is the most common form of primary glomerulonephritis. In IgAN, IgA1 molecules with incompletely galactosylated O-linked glycans in the hinge region (HR) are present in mesangial immunodeposits and in circulating immune complexes. It is not known whether the galactose deficiency in IgA1 proteins occurs randomly or preferentially at specific sites. We have previously demonstrated the first direct localization of multiple O-glycosylation sites on a single IgA1 myeloma protein by use of activated ion-electron capture dissociation (AI-ECD) Fourier transform ion cyclotron resonance (FT-ICR) tandem mass spectrometry. Here, we report the analysis of IgA1 O-glycan heterogeneity by use of FT-ICR MS and liquid chromatography FT-ICR MS to obtain unbiased accurate mass profiles of IgA1 HR glycopeptides from three different IgA1 myeloma proteins. Additionally, we report the first AI-ECD fragmentation on an individual IgA1 O-glycopeptide from an IgA1 HR preparation that is reproducible for each IgA1 myeloma protein. These results suggest that future analysis of IgA1 HR from IgAN patients and normal healthy controls should be feasible.

Synthesis of TNF- by mesangial cells cultured with polymeric anionic IgA role of MAPK and NF- B

Deposition of polymeric IgA1 (pIgA) in kidney mesangium is the hallmark of IgA nephropathy (IgAN). Current consensus is that a fraction of IgA1 molecules in the circulation of IgAN patients exhibit aberrant structures or properties that may lead to their deposition. Our previous findings suggest that the anionic property of IgA1 may play a role in mesangial IgA1 deposition in patients with IgAN. In the present study, the functional consequences of the binding of anionic polymeric IgA1 to human mesangial cells (HMCs) were investigated. Anionic polymeric IgA1 from IgAN patients and healthy subjects was isolated by sequential jacalin affinity chromatography, size exclusion chromatography using size exclusion and MonoQ ion exchange chromatography. HMCs were cultured with purified anionic polymeric IgA1 and the release of tumour necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) was examined by enzyme-linked immunosorbent assay. The signalling pathways involved in anionic pIgA-mediated HMC activation were examined by immunoblotting. Standard electrophoretic mobility shift assay (EMSA) was used to further examine whether the transcriptional factor NF-kappaB is associated in the signalling process. To define the mechanism of TNF-alpha and IL-6 production, HMCs were cultured with anionic pIgA in the presence or absence of p42/p44 mitogen-activated protein kinase (MAPK) inhibitor (PD98059), NF-kappaB inhibitor pyrrolidine dithiocarbamate (PDTC) or NF-kappaB blocking permeable peptides. Compared with less anionic pIgA or monomeric IgA1 (mIgA), anionic pIgA from patient with IgAN significantly increased cell proliferation (P<0.05) when cultured with HMC. These anionic pIgA significantly increased the synthesis of TNF-alpha (P<0.05) and IL-6 (P<0.05) in a dose and time-dependent manner. Furthermore, the increased synthesis of IL-6 and TNF-alpha by anionic pIgA in HMC was significantly diminished (P<0.01) in the presence of NF-kappaB inhibitor pyrrolidine dithiocarbamate and NF-kappaB blocking permeable peptides SN50 (P<0.01). The increased synthesis of IL-6 by anionic pIgA in HMC was reduced by inhibitor to NF-kappaB or p42/p44 MAPK and was abolished by the simultaneous presence of inhibitors to p42/p44 MAPK and NF-kappaB. The up-regulation of TNF-alpha was partially suppressed by inhibitor to NF-kappaB but not PD98059. Our results suggest that polymeric anionic IgA1 could activate HMC and increase the synthesis of TNF-alpha and IL-6. While both the p42/p44 MAPK and NF-kappaB pathways are essential in regulating the anionic pIgA-induced synthesis of IL-6, TNF-alpha synthesis mediated by anionic pIgA is partly dependent on NF-kappaB.

IgA nephropathy: what’s new?

Although IgA nephropathy has only been recognized as a definitive entity for fewer than 40 years, its place in the world as a prominent cause for progressive kidney disease is well established. The extent to which we understand the role of genetically derived abnormal forms of the IgA molecule in the disease is evolving, and this will, hopefully, translate into more specific modes of treatment for patients in the future. In the meantime, we have few specific therapeutic options, most of which have not been well studied in large numbers of patients. The extent to which we can define which patients are likely to progress--and hence should be considered candidates for treatment--will be discussed in this Commentary. In addition, the notion that some patients may have reached "the point of no return" will also be addressed. Unfortunately, most of the comments will be based on results obtained in studies conducted in adults--a situation that is very familiar to pediatric nephrologists.

Variants of theST6GALNAC2promoter influence transcriptional activity and contribute to genetic susceptibility to IgA nephropathy

IgA nephropathy (IgAN) is a polygenic disorder. Increasing evidence has implicated that aberrant glycosylation of IgA1 molecules, including alpha2,6 sialic acid defects, are involved in the pathogenesis of IgAN. In the present study, we designed an association study to investigate polymorphisms of two important genes, ST6GALNAC2 and NEU1, which are involved in the sialylation of the IgA1 molecule, in the susceptibility to IgAN. A total of 670 patients with histologically proven IgAN and 494 healthy controls were enrolled. Screening of SNPs in the coding and promoter regions of the ST6GALNAC2 and NEU1 genes was performed by sequencing. ST6-SNP1 (c.-988A>G), ST6-SNP2 [rs3840858:D>I(CGGC), c.-450_-449insCGGC], ST6-SNP3 (rs1867561:C>G, c.-135C>G), and ST6-SNP7 (rs2304921:G>A, c.186+12G>A) in the ST6GALNAC2 gene were selected as tagging SNPs. Functional evaluations of targets were assayed by luciferase activity. The alpha2,6 sialic acid contents of serum IgA1 in 497 patients were analyzed. Our results demonstrated that the frequency of haplotype ADG in the promoter region of ST6GALNAC2 was significantly higher in IgAN patients than that in controls (p=0.0069; odds ratio [OR]=1.36; 95% confidence interval [CI], 1.08-1.72). Furthermore, the ADG haplotype was associated with the deficient degrees of alpha2,6 sialic acid of IgA1 molecules in IgAN patients (r=0.408, p=0.0035). The ADG haplotype conferred significantly reduced promoter activity compared with the most common haplotype GDG in vitro (196.43+/-21.55 vs. 258.41+/-46.25; p=0.002). In the present study, we identified for the first time the ADG haplotype in the ST6GALNAC2 gene as a functional regulatory variant that may contribute to the genetic susceptibility in a subset of patients in whom the desialylation of IgA1 molecules was the main causative pathogenesis of IgAN.

Variants of C1GALT1 gene are associated with the genetic susceptibility to IgA nephropathy

IgA nephropathy (IgAN) is a polygenic disorder and the precise role of genetic factors remains elusive. Increasing evidences have implicated the aberrant galactosylation of IgA1 molecules in the pathogenesis of IgAN. The galactosyltransferase, core 1 beta3-Gal-T, and its chaperone, Cosmc, play important roles in beta1,3 glycosylation of IgA1 molecule. A case-control association study was performed to investigate the association between single-nucleotide polymorphisms (SNPs) of C1GALT1 and C1GALT1C1 genes and the susceptibility to IgAN. A total of 1164 subjects were enrolled, including 670 IgAN patients and 494 geographically matched healthy controls. Five SNPs, -734C/T, -465A/G, -330G/T, -292C/-, and 1365G/A in C1GALT1 were selected as tagging SNPs. The D allele and DD genotype of -292C/- in IgAN patients were significantly lower than in the controls (P<0.01). The frequency of haplotype YATIG (Y=C or T) was significantly lower in patients than in controls (0.0719 vs 0.1168, P=2.775 x 10(-4), odds ratio (OR)=0.70). The haplotype YAGDA (0.1236 vs 0.0791, P=3.815 x 10(-3), OR=1.77) and YATDG (0.0840 vs 0.0298, P=1.258 x 10(-5), OR=3.03) were significantly higher in patients than in controls. The present study suggested that the polymorphisms of C1GALT1 gene were associated with the genetic susceptibility to IgAN in Chinese population.

Development of Immunoglobulin A Nephropathy- Like Disease in β-1,4-Galactosyltransferase-I-Deficient Mice

Beta4 galactosylation of glycoproteins plays important roles in protein conformation, stability, transport, and clearance from the circulation. Recent studies have revealed that aberrant glycosylation causes various human diseases. Here we report that mice lacking beta-1,4-galactosyltransferase (beta4GalT)-I, which transfers galactose to the terminal N-acetylglucosamine of N- and O-linked glycans in a beta-1,4 linkage, spontaneously developed human immunoglobulin A nephropathy (IgAN)-like glomerular lesions with IgA deposition and expanded mesangial matrix. beta4GalT-I-deficient mice also showed high serum IgA levels with increased polymeric forms as in human IgAN. IgAN is the most common form of glomerulonephritis, and a significant proportion of patients progress to renal failure. However, pathological molecular mechanisms of IgAN are poorly understood. In humans, abnormal character of serum IgA, especially serum IgA1 with aberrant galactosylation and sialylation of O-glycans in its hinge region is thought to contribute to the pathogenesis of IgAN. Mouse IgA has N-glycans but not O-glycans, and beta4-galactosylation and sialylation of the N-glycans on the serum IgA from beta4GalT-I-deficient mice was completely absent. This is the first report demonstrating that genetic remodeling of protein glycosylation causes IgAN. We propose that carbohydrates of serum IgA are involved in the development of IgAN, whether the carbohydrates are O-glycans or N-glycans.

Sooty mangabey (Cercocebus torquatus atys) IGHG and IGHA genes

Antibodies are adaptor molecules of the immune system that link antigen recognition with the effector mechanisms responsible for antigen clearance. Several nonhuman primate species are widely used in biomedical research, especially for vaccine development and for AIDS-related studies. However, nonhuman primate antibody molecules have been characterized only partially and only in a few species. Here, we describe sooty mangabey (Cercocebus torquatus atys) IGHG and IGHA genes, which encode the heavy-chain constant region of IgG and IgA molecules, respectively. The four mangabey IGHG genes are highly homologous to the rhesus macaque and baboon IGHG genes (percent identity varies between 94.0 and 98.8, depending on the subclass), with most amino acid differences located in the hinge regions. Results obtained by real-time reverse transcription polymerase chain reaction show that the four IGHG genes are expressed at least at the mRNA level. The mangabey IGHA gene is highly homologous to the corresponding gene from rhesus macaques (percent identity ranges from 88.6 to 96.7, depending on the allele considered), the only other nonhominoid primate species for which the complete sequence of the IGHA gene is currently available. In the mangabey analyzed, two IGHA alleles are present, confirming that high levels of IGHA gene heterozygosity are present in monkey species. These results show that nonhuman primate gamma and alpha heavy chains differ from each other mostly at the level of the hinge region and that alpha sequence heterogeneity in nonhuman primate species is also present in other gamma regions. In addition, these results provide sequence information that can be used for residue frequency analysis of antibody heavy-chain constant region sequences.

Use of virus vectors for the expression in plants of active full‐length and single chain anti‐coronavirus antibodies

To extend the potential of antibodies and their derivatives to provide passive protection against enteric infections when supplied orally in crude plant extracts, we have expressed both a small immune protein (SIP) and a full‐length antibody in plants using two different plant virus vectors based on potato virus X (PVX) and cowpea mosaic virus (CPMV). The agr;SIP molecule consisted of a single chain antibody (scFv) specific for the porcine coronavirus, transmissible gastroenteritis virus (TGEV) linked to the α‐CH3 domain from human IgA. To express the full‐length IgA, the individual light and heavy chains from the TGEV‐specific mAb 6A.C3 were inserted into separate PVX constructs and plants were co‐infected with both constructs. Western blot analysis revealed the efficient expression of both the SIP and IgA molecules. Analysis of crude plant extracts revealed that both the plant‐expressed αSIP and IgA molecules could bind to and neutralize TGEV in tissue culture, indicating that active molecules were produced. Oral administration of crude extracts from antibody‐expressing plant tissue to 2‐day‐old piglets showed that both the αSIP and full‐length IgA molecules can provide in vivo protection against TGEV.

IgA-containing immune complexes in the urine of IgA nephropathy patients

Sera of IgA nephropathy (IgAN) patients contain high levels of circulating immune complexes composed of IgA1 molecules with aberrantly glycosylated hinge-region O-linked oligosaccharides and IgG or IgA1 antibodies with anti-glycan or anti-hinge-region peptide specificities. Due to damaged sieving properties of the glomerular capillary wall in IgAN, these immune complexes may appear in the urine. We collected urine samples from 29 patients with biopsy-proven IgAN (Group I), 27 proteinuric patients with non-IgA nephropathies (Group II) and 28 healthy volunteers (Group III). The levels of urinary IgA and IgG and IgA-IgG-containing immune complexes were measured by ELISA and standardized for urinary creatinine concentrations. The urinary IgA and IgG levels were significantly higher in Groups I and II than in Group III. Although the excretion of IgA as a fraction of total urinary protein was not significantly greater in IgAN patients than in patients with other renal diseases, the excretion of aberrantly glycosylated IgA1 was observed by western blot in 68% of the IgAN patients but in none of the healthy controls. The urinary levels of IgA-IgG immune complexes were significantly higher in Group I than in Groups II (P < 0.01) and III (P < 0.05). There was no significant difference in the levels between Groups II and III. These immune complexes had a molecular mass between 650-850 kDa, as shown by size-exclusion chromatography. The amounts of urinary IgA-IgG-containing immune complexes were significantly higher in patients with IgAN than in patients with non-IgA nephropathies or healthy controls.

Pathogenic IgA in IgA nephropathy: still the blind men and the elephant?

IgA nephropathy (IgAN), the most common glomerulonephritis worldwide, remains an important cause of end-stage renal failure. The pathology is characterized by mesangial deposition of IgA. The disease is now recognized as arising from anomalies of the IgA molecule and the kidneys are innocent bystanders. The immunochemical nature of the IgA molecule and its mesangial uptake command a pivotal role in the pathogenesis of IgAN.

To Treat or Not to Treat IgA Nephropathy? That Is the Question!

IgA nephropathy (IgAN) remains one of the most common forms of idiopathic glomerulonephritis in many populations worldwide (1,2). It also is a potentially serious condition in which from 4 to 38% of patients progress to ESRD by 10 yr (2). Given its frequency and significance, there should be a consensus on which treatment strategy is optimal for patients with IgAN (1,3,4). For those who work with patients with IgAN, the reasons for a lack of consensus are obvious. First, despite progress in understanding the mechanisms that are involved in IgAN, ranging from abnormalities of galactosylation of the hinge region of the IgA molecule to altered mesangial cell binding and incitement of inflammatory mediators, the pathogenesis of the disease remains unknown (1,5). Hence, specific treatment cannot be directed at the cause of the disease, and we are forced to use less specific disease modifiers. A second problem in dealing with the treatment of IgAN is the variability of glomerular manifestations and their severity. Some patients seem to show no progression of disease over many years, whereas others present with crescentic glomerulonephritis and progress …

Glycans on Secretory Component Participate in Innate Protection against Mucosal Pathogens

In mucosal secretions, secretory component (SC) is found either free or bound to polymeric IgA within the secretory IgA complex. SC displays numerous and various glycans, which are potential ligands for bacterial compounds. We first established that human SC (hSC) purified from colostrum (hSCcol) or produced in Chinese hamster ovary cells (hSCrec) exhibits the same lectin reactivity. Both forms bind to Clostridium difficile toxin A and functionally protect polarized Caco-2 cell monolayers from the cytopathic effect of the toxin. The interaction is mediated by glycans present on hSC and involves galactose and sialic acid residues. hSCcol and hSCrec were also shown to bind enteropathogenic Escherichia coli adhesin intimin and to inhibit its infectivity on HEp-2 cells in a glycan-dependent manner as well. SC remained operative in the context of the whole secretory IgA molecule and can therefore enhance its Fab-mediated neutralizing properties. On the contrary, hSC did not interact with three different strains of rotavirus (RF, RRV, and SA11). Accordingly, infection of target MA104 cells with these rotavirus strains was not reduced in the presence of either form of hSC tested. Although not a universal mechanism, these findings identify hSC as a microbial scavenger contributing to the antipathogenic arsenal that protects the body epithelial surfaces.

Binding capacity of in vitro deglycosylated IgA1 to human mesangial cells

IgA nephropathy (IgAN) is the most common glomerular disease and it is characterized by deposition of IgA1 molecules in mesangium. Recent studies had demonstrated that serum and mesangial IgA1 in IgAN were deglycosylated and IgA1 could bind to human mesangial cells (HMC) through a novel receptor. The aim of the current study is to investigate and compare the binding capacities of different in vitro deglycosylated IgA1 on human mesangial cells. Serum IgA1 was purified by jacalin affinity chromatography and then was desialylated (DesIgA1) and/or degalactosylated (Des/DeGalIgA1) with neuraminidase and/or β-galactosidase. The efficacy of deglycosylations was assessed by Peanut agglutinin (PNA) and Vicia villosa (VV) lectin. The sizes of normal IgA1 and deglycosylated IgA1 were determined by Sephacryl S-300 chromatography and binding capacities to primary HMC were evaluated by radioligand binding assays. Normal IgA1 and deglycosylated IgA1 could bind to HMC in a dose-dependent, saturable manner. The maximal binding capacities and binding sites/cell of DesIgA1 and Des/DeGalIgA were significantly higher than that of normal IgA1. However, more aggregated IgA1 was found in DesIgA1 and Des/DeGalIgA1. Scatchard analysis revealed a similar K d of normal IgA1 and deglycosylated IgA1. The current study suggested that the binding capacities of DesIgA1 and Des/DeGalIgA1 to HMC were significantly higher than that of normal IgA1, which at least in part was due to more macromolecular IgA1 in deglycoslated IgA1. However, there were no significant differences in the affinities of normal IgA1, DesIgA1 and Des/DeGalIgA1 with HMC. Deglycosylated IgA1 might play an important role in pathogenesis of IgAN.