Adsorptive filtration systems for effective removal of blood amyloid ß: a potential therapy for Alzheimer's disease.

Accumulation of amyloid-ß protein (Aß) in the brain causes cognitive impairment in Alzheimer's disease. We hypothesized that an extracorporeal system that rapidly removed Aß from the blood may accelerate Aß drainage from the brain. We previously reported that dialyzers remove blood Aßs effectively, mainly by adsorption on the inner surfaces of the hollow fibers, resulting in lower Aß accumulation in the brains of patients undergoing hemodialysis than the controls without hemodialysis. The aim of the present study was to create a more convenient and effective blood Aß removal system using adsorptive filtration, in which the filtrate returned to the body. Filtration from inside to outside of the fibers may enhance the adsorption of plasma Aßs on the surface of micropores inside the hollow fiber walls. Hence, pool solutions of 4 ng/mL synthetic Aß1-40 and Aß1-42 peptides (300 mL) or human plasma (1000 mL of 250-346 pg/mL Aß1-40 and 30-48 pg/mL Aß1-42) were circulated through polysulfone dialyzers at a flow rate of 50 mL/min to evaluate an adsorptive filtration system. The rates of Aß reduction from the pool solutions significantly increased along with the filtration rates. A filtration rate of > 1 mL/min, preferably 5-10 mL/min resulted in an 80-100% reduction of Aßs within 30 min of circulation. The rates of Aßs passing through the membrane walls were maintained around 0% for plasma Aßs during circulation. Thus, our adsorptive filtration systems may be useful for removing blood Aßs for patients with Alzheimer's disease.

Toward the treatment for Alzheimer's disease: adsorption is primary mechanism of removing amyloid ß protein with hollow-fiber dialyzers of the suitable materials, polysulfone and polymethyl methacrylate.

The accumulation of amyloid ß protein (Aß) in the brain reflects cognitive impairment in Alzheimer's disease. We hypothesized that the rapid removal of Aß from the blood by an extracorporeal system may act as a peripheral Aß sink from the brain. The present study aimed to determine the optimal materials and modality for Aß removal by hemodialyzers. In a batch analysis, hollow-fiber fragments of polysulfone (PSf) and polymethyl methacrylate (PMMA) showed greater removal efficiency of Aß than did other materials, such as cellulose-triacetates and ethylene-vinyl alcohol copolymer (PSf:PMMA at 30 min, 98.6 ± 2.4 %:97.8 ± 0.4 % for Aß1-40 and 96.6 ± 0.3 %:99.0 ± 1.0 % for Aß1-42). In a modality study, the Aß solution was applied to PSf dialyzers and circulated in the dialysis and Air-filled adsorption-mode (i.e., the outer space of the hollow fibers was filled with air) or phosphate-buffered saline (PBS)-filled adsorption modes. The Aß1-40 removal efficiency of the pre/post dialyzer in the Air-filled adsorption-mode was the highest (62.4 ± 12.6 %, p = 0.007). In a flow rate study in the Air-filled adsorption-mode, 200 mL/min showed the highest Aß1-40 reduction rate of pool solution (97.3 ± 0.8 % at 15 min) compared with 20 mL/min (p = 0.00001) and 50 mL/min (p = 0.00382). PMMA dialyzers showed similar high reduction rates. Thus, the optimal modality for Aß removal was the adsorption-mode with PSf or PMMA hollow fibers at around 50 mL/min flow rate, which seems to be suitable for clinical use.

A prospective study on blood Aß levels and the cognitive function of patients with hemodialysis: a potential therapeutic strategy for Alzheimer's disease.

To obtain the proof of concept of a novel therapy for Alzheimer's disease (AD), we conducted two prospective studies with hemodialysis patients who had amyloid ß protein (Aß) removed from their blood three times a week. One major pathological change in the brain associated with AD is Aß deposition, mainly 40 amino acids Aß1-40 and 42 amino acids Aß1-42. Impaired Aß clearance is proposed to be one cause of increased Aß in the AD brain. Thus, we hypothesized that an extracorporeal removal system of Aß from the blood may remove brain Aß and be a useful therapeutic strategy for AD. In the first prospective study, plasma Aß levels and the cognitive function of 30 hemodialysis patients (65-76 years old) were evaluated at baseline as well as 18 or 36 months after. Although plasma Aß1-40 levels either decreased or remained unchanged, levels of Aß1-42 either remained unchanged or increased at the second time point. Mini-Mental State Examination scores of most subjects increased or were maintained at the second time point. Aß1-40 influx into the blood correlated with MMSE at the second time point. In the second prospective study, five patients (51-84 years old) with renal failure were evaluated before and after the initiation of hemodialysis. Plasma Aß levels decreased, while cognitive function improved after initiating blood Aß removal. Therefore, long-term hemodialysis, which effectively removes blood Aß, might alter Aß influx and help maintain cognitive function.

Appropriate nonwoven filters effectively capture human peripheral blood cells and mesenchymal stem cells, which show enhanced production of growth factors.

Scaffolds, growth factors, and cells are three essential components in regenerative medicine. Nonwoven filters, which capture cells, provide a scaffold that localizes and concentrates cells near injured tissues. Further, the cells captured on the filters are expected to serve as a local supply of growth factors. In this study, we investigated the growth factors produced by cells captured on nonwoven filters. Nonwoven filters made of polyethylene terephthalate (PET), biodegradable polylactic acid (PLA), or chitin (1.2-22 µm fiber diameter) were cut out as 13 mm disks and placed into cell-capturing devices. Human mesenchymal stem cells derived from adipose tissues (h-ASCs) and peripheral blood cells (h-PBCs) were captured on the filter and cultured to evaluate growth factor production. The cell-capture rates strongly depended on the fiber diameter and the number of filter disks. Nonwoven filter disks were composed of PET or PLA fibers with fiber diameters of 1.2-1.8 µm captured over 70% of leukocytes or 90% of h-ASCs added. The production of vascular endothelial growth factor (VEGF), transforming growth factor ß1, and platelet-derived growth factor AB were significantly enhanced by the h-PBCs captured on PET or PLA filters. h-ASCs on PLA filters showed significantly enhanced production of VEGF. These enhancements varied with the combination of the nonwoven filter and cells. Because of the enhanced growth factor production, the proliferation of human fibroblasts increased in conditioned medium from h-PBCs on PET filters. This device consisting of nonwoven filters and cells should be investigated further for possible use in the regeneration of impaired tissues.

Comparison of effective impact among tonsillectomy alone, tonsillectomy combined with oral steroid and with steroid pulse therapy on long-term outcome of immunoglobulin A nephropathy.

BACKGROUND: To clarify the therapeutic impact of tonsillectomy and combined therapies of tonsillectomy plus steroid on the long-term prognosis of immunoglobulin A nephropathy (IgAN). METHODS: A retrospective study was conducted on 208 patients with IgAN between 1986 and 2009. According to the strategies for treatments, patients were divided into four groups: tonsillectomy and steroid pulse (TSP, n = 47), tonsillectomy and oral steroid (TOS, n = 33), tonsillectomy alone (T, n = 56), and N group (no particular therapy, n = 72). Multivariate analysis based on the Cox's regression model was used to assess the relative risk of reaching the outcome of doubling creatinine based on the influence of baseline prognostic factors. RESULTS: The mean observation periods were 53.8 months in the TSP group, 122.0 months in the TOS group, 102.9 months in the T group, and 84.6 months in the N group. During an observation period, serum creatinine levels doubled as follows: one in the TSP group (2.1 %), two in the TOS group (6.1 %), five in the T group (8.9 %), histological severity, and 22 in the N group (30.6 %). The Cox's regression proportional hazard model showed that gender, age, histological activity, dialysis induction risk and therapy were associated with doubling creatinine levels. Hazard ratios (95 % CI) and (P value) in T, TOS, and TSP groups versus N were 0.314 (0.11-0.93, P = 0.037), 0.213 (0.04-1.10, P = 0.065), and 0.032 (0.00-0.28, P = 0.002), respectively. CONCLUSION: A combination therapy of tonsillectomy and steroid pulse had the most significant therapeutic impact compared to other therapies.

A potential therapeutic system for Alzheimer's disease using adsorbents with alkyl ligands for removal of blood amyloid ß.

Amyloid beta proteins (Aß) in the brain are the main cause of Alzheimer's disease. Peripheral administration of Aß-binding substances, which may act as a sink for Aß from the brain, has been reported to reduce brain Aß. We previously found C16-cellulose beads had high Aß-removal activity in vitro. In this study, we investigated the optimum surface properties of adsorbents for removal of Aß in vitro and in humans. Batch analysis was performed with porous cellulose beads or silica beads with or without 2-22 methylene groups. Aß-removal activity of C16-cellulose beads increased with increasing alkyl chain length. In contrast, with cellulose the amount of Aß removed by the silica beads decreased with increasing alkyl chain length. Cellulose beads with 16 or 22 methylene groups were best (over 99 % removal) among all the beads tested (p ≤ 0.01). The adsorbent surfaces were analyzed by near-infrared spectroscopy, which revealed that the optimum beads had a sufficiently hydrophobic surface with an appropriate amount of adsorbed water accessible on the surface. Aß removal efficiency by C16-cellulose beads was investigated for 5 renal failure patients on hemodialysis, resulting in 51.1 ± 6.6 % for Aß1-40 and 43.8 ± 4.5 % for Aß1-42 (p ≤ 0.01). In conclusion, cellulose beads with 16 or 22 methylene groups and an appropriate amount of adsorbed water were the optimum Aß adsorbents. The device with C16-cellulose beads had high Aß removal activity in humans. These adsorbents might be useful for Alzheimer's disease therapy.

Potential therapeutic system for Alzheimer's disease: removal of blood Aßs by hemodialzyers and its effect on the cognitive functions of renal-failure patients.

The pathological changes of Alzheimer's disease include the deposition of amyloid ß protein (Aß) as senile plaques in the brain. We hypothesized that the rapid removal of Aßs from the blood may act as a peripheral Aß drainage sink from the brain. In this study, the plasma Aß concentrations and the cognitive functions were investigated for in 57 patients on hemodailysis (69.4 ± 3.8 years), 26 renal-failure patients without hemodialysis (66.6 ± 14.7 years), and 17 age-matched healthy controls (66.6 ± 4.1 years). The concentrations of plasma Aßs increased along with the decline of renal functions. Moreover, the renal-failure patients without hemodialysis and with poorer renal functions showed lower cognitive functions. The plasma concentrations of Aß(1-42) correlated with serum creatinine (P < 0.001) and Mini-Mental-State Examination scores (P = 0.017). The dialyzers effectively removed Aßs in the blood during hemodialysis sessions. The plasma Aß concentrations showed steady or slightly decreasing along with duration of hemodialysis. The total amount of Aßs removed during a hemodialysis session was calculated to be comparable to the Aßs dissolved in the blood and the cerebrospinal fluid. The MMSE scores of the hemodialysis patients showed no clear decrease in longer hemodialysis duration. Therefore, the therapeutic approach for Alzheimer's disease by removing Aßs from the blood is worthy of further investigation, including whether or not Aßs in the brain decrease.

Comparison of methods for profiling O-glycosylation: Human Proteome Organisation Human Disease Glycomics/Proteome Initiative multi-institutional study of IgA1.

The Human Proteome Organisation Human Disease Glycomics/Proteome Initiative recently coordinated a multi-institutional study that evaluated methodologies that are widely used for defining the N-glycan content in glycoproteins. The study convincingly endorsed mass spectrometry as the technique of choice for glycomic profiling in the discovery phase of diagnostic research. The present study reports the extension of the Human Disease Glycomics/Proteome Initiative's activities to an assessment of the methodologies currently used for O-glycan analysis. Three samples of IgA1 isolated from the serum of patients with multiple myeloma were distributed to 15 laboratories worldwide for O-glycomics analysis. A variety of mass spectrometric and chromatographic procedures representative of current methodologies were used. Similar to the previous N-glycan study, the results convincingly confirmed the pre-eminent performance of MS for O-glycan profiling. Two general strategies were found to give the most reliable data, namely direct MS analysis of mixtures of permethylated reduced glycans in the positive ion mode and analysis of native reduced glycans in the negative ion mode using LC-MS approaches. In addition, mass spectrometric methodologies to analyze O-glycopeptides were also successful.

PPAR-gamma agonist ameliorates kidney and liver disease in an orthologous rat model of human autosomal recessive polycystic kidney disease.

In autosomal recessive polycystic kidney disease (ARPKD), progressive enlargement of fluid-filled cysts is due to aberrant proliferation of tubule epithelial cells and transepithelial fluid secretion leading to extensive nephron loss and interstitial fibrosis. Congenital hepatic fibrosis associated with biliary cysts/dilatations is the most common extrarenal manifestation in ARPKD and can lead to massive liver enlargement. Peroxisome proliferator-activated receptor γ (PPAR-γ), a member of the ligand-dependent nuclear receptor superfamily, is expressed in a variety of tissues, including the kidneys and liver, and plays important roles in cell proliferation, fibrosis, and inflammation. In the current study, we determined that pioglitazone (PIO), a PPAR-γ agonist, decreases polycystic kidney and liver disease progression in the polycystic kidney rat, an orthologous model of human ARPKD. Daily treatment with 10 mg/kg PIO for 16 wk decreased kidney weight (% of body weight), renal cystic area, serum urea nitrogen, and the number of Ki67-, pERK1/2-, and pS6-positive cells in the kidney. There was also a decrease in liver weight (% of body weight), liver cystic area, fibrotic index, and the number of Ki67-, pERK1/2-, pERK5-, and TGF-ß-positive cells in the liver. Taken together, these data suggest that PIO inhibits the progression of polycystic kidney and liver disease in a model of human ARPKD by inhibiting cell proliferation and fibrosis. These findings suggest that PPAR-γ agonists may have therapeutic value in the treatment of the renal and hepatic manifestations of ARPKD.

Differential expression of glycogenes in tonsillar B lymphocytes in association with proteinuria and renal dysfunction in IgA nephropathy.

Aberrant O-glycosylation of serum and tonsillar IgA1 is one of the main pathogeneses of IgA nephropathy (IgAN). However, the synthesis of underglycosylated IgA1 in tonsils has not yet been characterized. This study examined tonsillar B lymphocytes of IgAN (n=34) using tonsils derived from patients with chronic tonsillitis (n=24) and sleep apnea syndrome (n=14) as a control. Gene expression of beta1,3-galactosyltransferase (beta3GalT), and the core 1 beta3GalT-specific molecular chaperone, Cosmc, UDP-N-acetyl-alpha-D-galactosamine: polypeptide N-acetylgalactosaminyl-transferase 2, were significantly decreased in tonsillar CD19-positive B lymphocytes from IgAN patients compared to control tonsillar tissues as determined by real-time RT-PCR. Tonsillar B cell beta3GalT gene expression significantly correlated with estimated GFR and negatively correlated with proteinuria and histological injury score. Western blotting showed the protein expression of beta3GalT in the tonsils to significantly decrease in IgAN in comparison to the controls. These data suggest the downregulation of beta3GalT in tonsillar B lymphocytes to be closely associated with the clinical characteristics of IgAN.

Evaluation of the specific structures of IgA1 hinge glycopeptide in 30 IgA nephropathy patients by mass spectrometry.

BACKGROUND: Although many reports have described the abnormal structures of O-glycan in the IgA1 hinge region in IgA nephropathy (IgAN), the specific glycopeptide that can be used as a diagnostic biomarker for IgAN has not yet been identified. To pursue this diagnostic approach, we used mass spectrometric analysis to search for specific structures in IgA1 hinge glycopeptides in 30 IgAN patients contrasted with 30 healthy controls. METHOD: IgA1 hinge glycopeptides were individually purified from the sera of 30 biopsy-proven IgAN patients and 30 healthy controls. The structure of each glycopeptide was analyzed by ion trap mass spectrometry. Sugar conformations in each glycopeptide were estimated by collision-induced dissociation tandem mass spectrometry. Furthermore, to search for specific O-glycans in IgAN patients with a statistical significance, the identified hinge glycopeptides were analyzed by Fisher exact test. RESULT: A total of 57 hinge glycopeptides were identified from each of the 2 sample groups. Among the structures of the hinge glycopeptides identified, statistical significance was found for 6 glycopeptides (O-glycan compositions were 33-mer hinge core peptides + xGalNAc + yGal + zNeu5Ac; x:y:z = 5:3:3, 5:3:0, 5:2:1, 4:2:2, 3:1:1, 3:1:0) by Fisher exact test (p<0.05). In these 6 O-glycan compositions, 3 compositions (x:y:z = 4:2:2, 3:1:1, 3:1:0) were only observed in IgAN patients and were absent in the 30 controls. CONCLUSION: Statistically specific O-glycans were identified in 30 IgAN patients compared with 30 controls. These results open the possibility for preparation of lectin and/or antibodies binding to specific glycopeptides in IgAN.

Removal of Aß Oligomers from the Blood: A Potential Therapeutic System for Alzheimer's Disease.

PURPOSE: Amyloid-ß protein (Aß) is one of the causative proteins of Alzheimer's disease. We have been developing extracorporeal blood Aß-removal systems as a method for enhancing Aß clearance from the brain. We reported previously that medical adsorbents and hemodialyzers removed Aß monomers from peripheral blood, which was associated with influx of Aß monomers from the brain into the bloodstream. Our intent here was to develop a method to promote clearance of Aß oligomers and to provide an estimate of the molecular size of intact Aß oligomers in plasma. METHODS: Two hollow-fiber devices with different pore sizes (Membranes A and B) were evaluated as removers of Aß oligomers with human plasma in vitro. The concomitant removal of Aß oligomers and monomers was investigated by using Membrane B and hexadecyl alkylated cellulose beads or polysulfone hemodialyzers. Double-filtration plasmapheresis with Membrane A was investigated as an approach for the removal of plasma Aß oligomers in humans. RESULTS: Aß oligomers were effectively removed by both Membranes A and B. The increase of Aß oligomers in plasma was observed just after the removal of plasma Aß oligomers in humans. The intact molecular size of major Aß oligomers in the plasma was estimated to be larger than albumin at approximately 60 kDa or more. Additionally, the concomitant removal of Aß monomers and oligomers evoked dissociation of larger Aß oligomers into smaller ones and monomers. CONCLUSION: Aß oligomers were cleared from plasma both in vitro and in human subjects by using hollow-fiber membranes with large pores, indicating that their intact sizes were mostly larger than 60 kDa. Aß oligomers in peripheral circulation were increased after some clearances in human. Further investigation will determine whether the Aß oligomers detected in circulation after clearance were via influx from the brain.

Expression of tumor necrosis factor receptors on granulocytes in patients with myeloperoxidase anti-neutrophil cytoplasmic autoantibody-associated vasculitis.

BACKGROUND/AIMS: To clarify the clinical significance of tumor necrosis factor (TNF) receptors in patients with myeloperoxidase (MPO)-anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis, we evaluated the cell surface expression of TNF receptor 1 (TNFR1) and TNF receptor 2 (TNFR2). PATIENTS AND METHODS: 43 patients with MPO-ANCA-associated vasculitis, 16 patients with chronic renal failure, 10 patients with sepsis, 15 patients with systemic lupus erythematosus, and 18 healthy controls were enrolled in this study, and the surface expression levels of TNFR1, TNFR2, CD63, and CD64 on granulocytes were assessed. In 21 patients with MPO-ANCA-associated vasculitis, soluble TNFR1 (sTNFR1), soluble TNFR2(sTNFR2), and TNF-alpha in the serum were also measured. RESULTS: The surface expression levels of TNFR1 and TNFR2 on granulocytes were significantly higher in patients with MPO-ANCA-associated vasculitis than in the healthy controls, and positively correlated with the Birmingham Vasculitis Activity Score (BVAS). The levels of sTNFR1, sTNFR2, and TNF-alpha in the serum were also significantly higher in patients with MPO-ANCA-associated vasculitis than in the healthy controls. Serum levels of sTNFR1 and sTNFR2 correlated with serum creatinine, while the surface expression of TNFR1 and TNFR2 on the granulocytes did not. There was no significant correlation between the BVAS and CD63 or BVAS and CD64. CONCLUSION: The surface expression levels of TNFR1 and TNFR2 on granulocytes were upregulated in patients with MPO-ANCA-associated vasculitis and reflected disease activity.

O-linked oligosaccharides of the IgA1 hinge region: roles of its aberrant structure in the occurrence and/or progression of IgA nephropathy.

Primary IgA nephropathy (IgAN) has been regarded as an immune complex-mediated glomerulonephritis characterized immunohistologically by the predominant deposition of IgA in the glomerular mesangial area with a variety of histopathologic injuries (Clarkson et al. in Ann Rev Med 38:157-168, 1987). In 1992, the characteristic structure of O-linked oligosaccharides (O-glycans) in the IgA1 hinge and its possible aberrancy were simultaneously and independently proposed by Mesteckey et al. (Cont Nephrol 104:172-182, 1993), and our group (Cont Nephrol 104:217, 1993) at the International Congress of Nephrology (IgA Nephropathy 25th year) held in Nancy, France. Since then, the aberrancy has been confirmed by several research groups and is suspected to play a role in the occurrence and/or the progression of IgAN. At the end of the 1980s, I took an interest in the existence of O-glycans in the hinge region of IgA1 and have pursued the structure of the carbohydrate chains. Since an excellent review on the structure and the role of the carbohydrate in IgA molecules was recently published by Narita et al. (Clin Ex Nephrol 12:332-338, 2008), this review focuses on the process by which I developed the idea of aberrant O-glycosylation in IgA1 molecules in IgAN patients and summarizes our recent observations concerning IgA1 molecules.

Serum levels of 5-s-cysteinyldopa are correlated with skin colors in hemodialysis patients but not in peritoneal dialysis patients.

BACKGROUND: Diffuse hyperpigmentation is common in patients with chronic renal failure undergoing hemodialysis (HD) or peritoneal dialysis (PD). We previously reported that serum levels of 5-S-cysteinyldopa (5SCD, a pheomelanin precursor) and pheomelanin were significantly elevated in HD patients. METHODS: Skin color was assessed using a Mexameter that measures the melanin index (MI) and the erythema index (EI). The upper inner arms (non-sun-exposed site) and the foreheads (sun-exposed site) of HD and PD patients and control subjects were analyzed. RESULTS: MI values on the upper inner arms and on the foreheads of HD and PD patients were significantly higher than in controls. In HD patients, significant correlations were found for serum 5SCD levels with MI and EI on the upper inner arm, and for EI on the forehead. In PD patients, no such correlations were found. CONCLUSIONS: Hyperpigmentation in HD patients results partly from accumulation of pheomelanin in the skin.

Influx of Tau and Amyloid-ß Proteins into the Blood During Hemodialysis as a Therapeutic Extracorporeal Blood Amyloid-ß Removal System for Alzheimer's Disease.

The accumulation of amyloid-ß protein (Aß) and tau in the brain is a major pathological change related to Alzheimer's disease. We have continued to develop Extracorporeal Blood Aß Removal Systems (E-BARS) as a method for enhancing Aß clearance from the brain. Our previous report revealed that dialyzers effectively remove blood Aß and evoke large Aß influxes into the blood, resulting in a decrease in brain Aß accumulation after initiating hemodialysis, and that patients who underwent hemodialysis had lower brain Aß accumulation than those who did not. Here, plasma total tau concentrations from 30 patients undergoing hemodialysis were measured using an ultrasensitive immunoassay and compared to those from 11 age-matched controls. Plasma total tau concentrations were higher in patients with renal failure regardless of whether they underwent hemodialysis, suggesting the involvement of the kidneys in tau degradation and excretion. Hemodialyzers effectively removed blood Aß but not extracorporeal blood tau. The influx of tau into the blood was observed at around the 1 h period during hemodialysis sessions. However, the influx amount of tau was far smaller than that of Aß. Furthermore, histopathological analysis revealed similar, not significantly less, cerebral cortex phosphorylated tau accumulation between the 17 patients who underwent hemodialysis and the 16 age-matched subjects who did not, although both groups showed sparse accumulation. These findings suggest that hemodialysis may induce both tau and Aß migration into the blood. However, as a therapeutic strategy for Alzheimer's disease, it may only be effective for removing Aß from the brain.

Serum under-galactosylated IgA1 is increased in Japanese patients with IgA nephropathy.

BACKGROUND: Immunoglobulin A nephropathy (IgAN) is characterized by an aberrant structure of O-glycans in the IgA1 hinge region. Recently, under-galactosylated IgA1 has been found to be increased in Caucasian IgAN patients. Thus, we examined this in Japanese IgAN patients. METHODS: An enzyme-linked immunosorbent assay of binding between Helix aspersa (HAA) and serum IgA was performed in Japanese IgAN patients and the HAA-IgA binding levels were compared among IgAN patients (n = 41), patients with other forms of kidney disease (OKD, n = 43) and healthy controls (n = 38). The clinicopathological severity of IgAN was then analysed between patients with high and low HAA-IgA binding levels. The levels were also compared in 11 patients before and after the combination of tonsillectomy and steroid pulse therapy. Furthermore, we examined the O-glycan structure of IgA1 hinge glycopeptides by mass spectrometry (MS). RESULTS: The HAA-IgA binding levels were significantly higher in IgAN patients compared with either healthy controls (P = 0.0025) or those with OKD (P = 0.016). To reflect the absolute level of under-galactosylated IgA, we multiplied the HAA-IgA binding level by the serum IgA concentration to produce an indicative value. The specificity and sensitivity of this value were 89% and 66%, respectively. MS showed that peak distribution of IgA1 hinge glycopeptides was shifted to smaller molecular weights in high HAA-IgA-binding IgAN patients. There was no correlation between the HAA-IgA binding level and either disease severity or the use of combination therapy. CONCLUSIONS: HAA-IgA binding is significantly increased in Japanese IgAN patients. This potential IgAN marker is not affected by disease severity or therapeutic intervention.

The HLA-DRB1*1501 allele is prevalent among Japanese patients with anti-glomerular basement membrane antibody-mediated disease.

BACKGROUND: We aimed to clarify the relationship between HLA-DRB1(*)1501 and anti-glomerular basement membrane (GBM) antibody-mediated disease in Japanese patients. MATERIALS: Samples were collected from 16 anti-GBM antibody-positive patients who were admitted to our department or related hospitals from December 1990 to October 2005. We analysed clinical and laboratory data, kidney biopsy findings, and the HLA-DR phenotypes and HLA-DRB1 alleles of the patients. RESULTS: Among the 16 patients, 15 had HLA-DR15 [the phenotype frequency (PF) was 93.8%], 7 were positive for DR4 (the PF was 43.8%) and 5 were positive for DR9 (the PF was 31.3%). The allele frequency of HLA-DRB1(*)1501 was 46.4% (13/28), which was significantly different from Japanese controls (11.6%) (P < 0.001). In contrast, the frequency of HLA-DRB1(*)1502 was not different from controls (0/28). The odds ratio of HLA-DRB1(*)1501 in these patients was 6.4 (95% CI: 2.4-16.5). CONCLUSION: The present study demonstrated that Japanese patients with anti-GBM antibody-mediated disease are very likely to carry the HLA-DRB1(*)1501 but not the HLA-DRB1(*)1502 allele.

Similarities between N-glycan glycoform of tonsillar IgA1 and that of aberrant IgA1 abundant in IgA nephropathy patient serum.

BACKGROUND: There are many reports on the presence of an incompletely glycosylated O-linked oligosaccharide(s) in the IgA1 hinge region of some IgA nephropathy patients. As the candidates of such IgA1, tonsillar IgA1 and aberrant IgA1, which are abundant in an IgA nephropathy patient, were proposed. On the other hand, in mice, the abnormality of the N-linked oligosaccharide chain of IgA induced the IgA nephropathy. Therefore, analyses of the N-glycan glycoform on serum IgA1, aberrant IgA1 and tonsillar IgA1 were carried out using the 3-dimensional mapping method. RESULTS: The sugar chain composition was almost the same in these 3 IgA1 preparations. However, the structural characteristics for the aberrant IgA1 showed a drastic increase in the neutral N-glycans; in particular, 25% of the sugar chains in the aberrant IgA1 were the high mannose-type as compared with approximately 5%-6% in the serum IgA1 and tonsillar IgA1. The neutral complex-type N-glycan chain with fucose was higher in both the aberrant IgA1 and tonsillar IgA1 than in the serum IgA1. A typical component in the aberrant IgA1 was the fully galactosylated biantenna with the fucose residue. CONCLUSIONS: We found an abnormality in the N-linked oligosaccharides of the aberrant IgA1. In addition to our previous report about the abundance of asialo-O-linked oligosaccharide in both the tonsillar IgA and aberrant IgA, our results concerning the N-glycan glycoform of the aberrant IgA showed the possible promotion of its self-aggregation and its glomerular deposition by the synergistic difference in the O- and N-linked carbohydrate chains, and also the derivation of the aberrant IgA1 in the sera from the tonsillar tissue.

Effects of cytapheresis on tumor necrosis factor receptor and on expression of CD63 in myeloperoxidase--antineutrophil cytoplasmic autoantibody-associated vasculitis.

To evaluate the therapeutic potential of cytapheresis in myeloperoxidase-antineutrophil cytoplasmic autoantibody (MPO-ANCA)-associated vasculitis, plasma levels of soluble tumor necrosis factor receptors (sTNFR1, sTNFR2) and the expression of TNFR1, TNFR2, and CD63 on granulocytes were measured. The levels of sTNFR1 and sTNFR2, and the expression of TNFR1 and TNFR2 were significantly higher in MPO-ANCA-associated vasculitis patients than in normal controls. The levels of sTNFR1 and sTNFR2 increased significantly after cytapheresis (P < 0.001). The expression of TNFR1 showed a tendency to decrease after cytapheresis (P = 0.0535). The expression of CD63 decreased significantly after cytapheresis (P < 0.05). Because sTNFR1 and sTNFR2 act as TNF-antagonists, the increases of sTNFR1 and sTNFR2 after cytapheresis might contribute to inhibit the action of TNF-alpha. The decreased expression of TNFR1, which mediates the signal for polymorphonuclear cell respiratory burst, might also contribute to the reduction of inflammation. From these results, the inhibition of TNF action and removal of degranulated granulocytes appear to be related to the mechanism whereby cytapheresis can exert a beneficial and therapeutic function in the treatment of MPO-ANCA-associated vasculitis.