IgA Nephropathy Group Research Articles

Increased phosphatidylserine-exposing microparticles and their originating cells are associated with the coagulation process in patients with IgA nephropathy.

Relatively little information is available about phosphatidylserine positive (PS(+)) microparticles (MPs) and their originating cells in IgA nephropathy (IgAN) despite well-established intraglomerular coagulation. Our objectives were to detect PS exposure on MP membranes and MP-origin cells and to evaluate its role in procoagulant activity (PCA) and fibrin formation and their association with pathological lesions in the disease. Patients with IgAN and healthy controls were studied. Lactadherin was used to quantify PS exposure on MPs and MP-origin cells. PCA of MPs and MP-origin cells was evaluated by clotting time and purified coagulation complex assays. Fibrin production was determined by turbidity. PS exposure, fibrin strands and FVa/Xa binding were observed on MPs/cells using confocal microscopy. Using flow cytometry, we found that IgAN patients had high levels of PS(+) MPs derived from lymphocytes, monocytes, neutrophils, platelets, erythrocytes and endothelial cells (ECs). The PS exposure on MP-origin cells also increased in these patients. MPs and MP-origin cells (leukocytes, platelets and erythrocytes) isolated from IgAN patients and ECs cultured with IgAN serum had a significantly shorter median coagulation time (P < 0.001), higher median intrinsic FXa (P < 0.001) and higher thrombin (P < 0.001) generation than controls. These coagulation functional assays were associated with the glomerular lesions. The lesions were also correlated with glomerular fibrin deposition (all P < 0.05). In the presence of patient MPs or their related cells, fibrin formation peaked faster with a higher maximum turbidity when compared with healthy controls. Blocking PS with lactadherin in the IgAN group prolonged coagulation time to control levels, inhibited the PCA up to 80% and markedly reduced fibrin formation. More importantly, we observed that fibrin strands formed on MPs and ECs in the same regions that bound lactadherin, similar to the FVa/Xa costaining. We find that high levels of PS(+) MPs and the MP-origin cells are associated with the coagulation process in IgAN, and this may provide a previously unrecognized contribution to intraglomerular coagulation.

Efficacy of triptolide on the apoptosis of tonsillar mononuclear cells from patients with IgA nephropathy

Aims: This study aimed to evaluate the extent of apoptosis of tonsillar mononuclear cells (TMCs) derived from patients with IgA nephropathy (IgAN) and the effects of triptolide (TP) on the apoptosis of these TMCs. Methods: TMCs were isolated from tonsillar tissues of patients with IgAN or chronic tonsillitis (control group). Rates of TMCs apoptosis were measured by annexin V-fluorescein isocyanate (FITC)/propidium iodide (PI)-labeled flow cytometry (FCM). Expression levels of Bcl-2 family proteins were quantified by immunohistochemistry of fixed tonsillar sections and Western blot analyzes of TMCs lysates. TMCs from IgAN patients were treated 10, 20, or 30 ng/mL TP for 24 h and then evaluated for viability by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, for the percentage of apoptotic cells by FCM, and for the relative expression levels of Bcl-2 family proteins by Western blot analysis. Results: Compared to TMCs from the control group, TMCs from the IgAN group demonstrated lower rates of apoptosis, higher expression levels of the anti-apoptosis proteins Bcl-2 and Bcl-xL, and lower expression levels of the pro-apoptosis protein Bax. Treatment of IgAN patient-derived TMCs with 10, 20, or 30 ng/mL TP for 24 h suppressed the viability and promoted the apoptosis of TMCs in a dose-dependent manner. Western blot analysis revealed a TP dose-dependent decrease in Bcl-2 and Bcl-xL expression levels, in parallel with increased Bax protein levels. Conclusion: TMCs from IgAN patients may be in a state of inhibited apoptosis mediated by Bcl-2 family proteins, which may be reversed by TP treatment.

Retrospective study of primary IgA nephropathy with crescent formation and/or rapidly progressive glomerulonephritis in children

IgA nephropathy is the most common type of glomerulonephritis in the world. Its clinical and pathological manifestations vary. A few of the patients with IgA nephropathy present with rapidly progressive glomerulonephritis (RPGN) and/or crescent formation. Their conditions are serious and acute, but there are few reports on their characteristics, treatment and outcome. This study aimed to analyze the clinicalopathological features, treatment and prognosis of primary IgA nephropathy in children, to provide a reference for clinical diagnosis and treatment. A retrospective study was conducted in children with primary IgA nephropathy with crescent formation and/or rapidly progressive glomerulonephritis admitted to our department from 2000 to 2014. The patients meeting the inclusion and exclusion criteria were included. Patients were divided into RPGN group and non-RPGN group according to the clinical manifestations, crescent formation group and non-crescent group, crescentic IgA nephropathy group and non-crescentic IgA nephropathy group according to renal biopsy. Their clinical manifestations and pathological features, treatment and prognosis were compared. A total of 265 patients were recruited, 10 patients (3.8%) had RPGN, 151 patients (57.0%) had crescent formation, 19 cases (7.2%) showed crescentic IgA nephropathy.Compared with non-RPGN group, RPGN group showed more gross hematuria, higher serum creatinine, lower creatinine clearance correction at biopsy and follow-up, and more crescentic IgA nephropathy (P<0.05). The percent of patients who received methylprednisolone pulse and blood purification therapy in RPGN group is higher than that of non-RPGN group (P<0.05). Compared with non-crescent group, crescent formation group showed more gross hematuria at biopsy and follow-up, higher serum creatinine at biopsy, lower creatinine clearance correction, more 24-hour urinary protein at biopsy and higher serum creatinine at follow-up (P<0.05). The percentage of patients received more methylprednisolone pulse, oral steroids, cyclophosphamide pulse in crescent formation group was higher than that of non-crescent group (P<0.05). Compared with non-crescentic IgA nephropathy group, crescentic IgA nephropathy group showed more RPGN percent, higher serum creatinine, more 24-hour urinary protein at biopsy (P<0.05). The percentage of patients who received more methylprednisolone pulse and blood purification therapy in crescentic IgA nephropathy group was more than non-crescentic IgA nephropathy group (P<0.05). At follow-up, 20.0% of the patients with RPGN and crescent nephritis returned to normal renal function and the percent of crescent glomerulonephritis but not RPGN was 71.4%, RPGN but not crescent glomerulonephritis was 80.0%, crescent formation but not crescent nephritis was 87.5%. In primary IgA nephropathy with crescent formation and/or rapidly progressive glomerulonephritis, the patients with both RPGN and crescentic IgA nephropathy showed the worst clinical manifestations, its prognosis was worst while the patients with crescent formation showed the mildest clinical manifestations and best prognosis.

Increased frequencies of memory and activated B cells and follicular helper T cells are positively associated with high levels of activation‑induced cytidine deaminase in patients with immunoglobulin A nephropathy.

The aim of the present study was to examine the frequencies of different subsets of B and follicular helper T (Tfh) lymphocytes in patients with immunoglobulin A nephropathy (IgAN), and investigate the potential underlying mechanism. A total of 27 patients with IgAN and 10 healthy controls (HC) were recruited for analysis of the frequencies of different subsets of B and Tfh cells. ELISA was used to analyze the concentration of serum interleukin (IL)‑21. The transcriptional levels of activation‑induced cytidine deaminase (AID) in the B cells were determined using reverse transcription‑quantitative polymerase chain reaction, while the translational levels of AID were analyzed using western blotting. The frequencies of circulating memory and activated B cells and Tfh cells were found to be significantly increased in the IgAN groups, compared with those of the HC group, although the number of plasma cells were not significantly different between the two IgAN groups. In addition, the serum levels of IL‑21 were found to be higher in the patients with IgAN, and correlated with 24‑h proteinuria. IL‑21 also enhanced the expression levels of AID in the B cells. The data of the present study revealed that the high levels of memory and activated B cells and Tfh cells were positively associated with the progression of IgAN, and that this may be mediated by the overexpression of AID, which is potentially regulated by IL‑21.

Profiling and initial validation of urinary microRNAs as biomarkers in IgA nephropathy.

Background. MicroRNAs (miRNAs) have been found in virtually all body fluids and used successfully as biomarkers for various diseases. Evidence indicates that miRNAs have important roles in IgA nephropathy (IgAN), a major cause of renal failure. In this study, we looked for differentially expressed miRNAs in IgAN and further evaluated the correlations between candidate miRNAs and the severity of IgAN.Methods. Microarray and RT-qRCR (real-time quantitative polymerase chain reaction) were sequentially used to screen and further verify miRNA expression profiles in urinary sediments of IgAN patients in two independent cohorts. The screening cohort consisted of 32 urine samples from 18 patients with IgAN, 4 patients with MN (membranous nephropathy), 4 patients with MCD (minimal changes disease) and 6 healthy subjects; the validation cohort consisted of 102 IgAN patients, 41 MN patients, 27 MCD patients and 34 healthy subjects. The renal pathological lesions of patients with IgAN were evaluated according to Lee’s grading system and Oxford classification.Results. At the screening phase, significance analysis of microarrays analysis showed that no miRNA was differentially expressed in the IgAN group compared to all control groups. But IgAN grade I–II and III subgroups (according to Lee’s grading system) shared dysregulation of two miRNAs (miR-3613-3p and miR-4668-5p). At the validation phase, RT-qPCR results showed that urinary level of miR-3613-3p was significantly lower in IgAN than that in MN, MCD and healthy controls (0.47, 0.44 and 0.24 folds, respectively, all P < 0.01 by Mann–Whitney U test); urinary level of miR-4668-5p was also significantly lower in IgAN than that in healthy controls (0.49 fold, P < 0.01). Significant correlations were found between urinary levels of miR-3613-3p with 24-hour urinary protein excretion (Spearman r = 0.50, P = 0.034), eGFR (estimated glomerular filtration rate) (r = − 0.48, P = 0.043) and Lee’s grades (r = 0.57, P = 0.014). Similarly, miR-4668-5p was significantly correlated with eGFR (r = − 0.50, P = 0.034) and Lee’s grades (r = 0.57, P = 0.013). For segmental glomerulosclerosis according to Oxford classification, patients scored as S0 had significantly lower levels of urinary miR-3613-3p and miR-4668-5p than those scored as S1 (0.41 and 0.43 folds, respectively, all P < 0.05).Conclusions. The expression profile of miRNAs was significantly altered in urinary sediments from patients with IgAN. Urinary expression of miR-3613-3p was down-regulated in patients with IgAN. Moreover, urinary levels of both miR-3613-3p and miR-4668-5p were correlated with disease severity. Further studies are needed to explore the roles of miR-3613-3p and miR-4668-5p in the pathogenesis and progression of IgA nephropathy.

Study of correlation between polymorphism of ST6GALNAC2 and susceptibility to IgA nephropathy.

The aim of the present study was to explore the correlation between single nucleotide polymorphisms (SNPs) rs3840858 and rs2304921 in a specific α-2,6 sialyltransferase gene, ST6GALNAC2, and the susceptibility to immunoglobulin (IgA) nephropathy (IgAN). The distributions of genotypes of SNPs rs3840858 and rs2304921 in ST6GALNAC2 were detected by direct sequencing. The distributions of the genotype and allele frequencies of rs3840858 in patients with IgAN were significantly different from those in the control group (genotypes, P=0.001; alleles, P=0.001). The DI genotype ratio (17.8%) in the IgAN group was higher than that in the control group (5.6%) and the I allele frequency (8.9%) in the IgAN group was higher than that in the control group (2.8%). Univariate logistic regression analysis indicated that rs3840858 polymorphism is a risk factor of IgAN (P=0.001). The risk of developing IgAN in individuals who carried the DI genotype was 3-fold higher than that in individuals who carried the DD genotype [odds ratio (OR)=3.676, 95% confidence interval (CI)=1.284-10.519], and the risk of developing IgAN in individuals who carried the I allele was higher than that in individuals who carried the D allele (OR=3.415, 95% CI=1.223-9.531). The distributions of the genotype (AA, AG and GG) and allele (A and G) frequencies of rs2304921 did not have a statistically significant difference between patients with IgAN and those without (P>0.05). The SNP rs3840858 in the ST6GALNAC2 gene may be associated with the risk of developing IgAN in the population studied; however, polymorphism of rs2304921 appears to be irrelevant to the risk of developing IgAN in this population.

Renal biopsy findings and clinical indicators of patients with hematuria without overt proteinuria.

BackgroundWhether to perform a renal biopsy for isolated hematuria remains a matter of controversy. We performed renal biopsy in hematuria without overt proteinuria patients and reported the proportion of glomerulonephritis, pathological activities, and statistical analysis of indicators associated with glomerulonephritis.MethodsAmong 203 patients who underwent renal biopsy in Okubo Hospital, Japan, between January 2008 and October 2013, we identified 56 patients who fulfilled the criteria: (1) urine dipstick examination shows equal to or greater than ± blood on three or more visits, (2) proteinuria <0.3 g/day (g/g Cr), (3) eGFR ≧60 ml/min/1.73 m2, and (4) no current medication for renal disease. We investigated biopsy findings and compared the clinical indicators in the IgA nephropathy (IgAN) and non-IgAN group.ResultsThe pathological diagnosis was IgAN in 35 cases (62 %), thin basement membrane disease (TBMD) in 7 (13 %), minor glomerular abnormality (MGA) in 6 (11 %), glomerular basement membrane (GBM) abnormality in 5 (9 %), and others in 3 (5 %). The histological grade of IgAN was I in 90 % and II in 10; 31 % of patients had some crescentic lesions. Comparisons between the IgAN and non-IgAN group revealed significant differences in age of onset (26 ± 13 vs. 34 ± 17 years, p = 0.04), serum IgA (340 ± 114 vs. 220 ± 101 mg/dl, p < 0.01), proteinuria (0.08 [0–0.25] vs. 0 [0–0.23] g/day [g/gCr], p < 0.01), and the presence of poikilocytes (40 vs. 10 %, p = 0.02).ConclusionsThe proportion of IgAN in hematuria without overt proteinuria was high and the pathological activities were variable. Patients with hematuria without overt proteinuria should continue their medical follow-up and the best timing of biopsy may be controversial for these patients who have multiple risk factors of IgAN.

DNA Methylation in Cosmc Promoter Region and Aberrantly Glycosylated IgA1 Associated with Pediatric IgA Nephropathy

IgA nephropathy (IgAN) is one of the most common glomerular diseases leading to end-stage renal failure. Elevation of aberrantly glycosylated IgA1 is a key feature of it. The expression of the specific molecular chaperone of core1ß1, 3galactosyl transferase (Cosmc) is known to be reduced in IgAN. We aimed to investigate whether the methylation of CpG islands of Cosmc gene promoter region could act as a possible mechanism responsible for down-regulation of Cosmc and related higher secretion of aberrantly glycosylated IgA1in lymphocytes from children with IgA nephropathy. Three groups were included: IgAN children (n = 26), other renal diseases (n = 11) and healthy children (n = 13). B-lymphocytes were isolated and cultured, treated or not with IL-4 or 5-Aza-2’-deoxycytidine (AZA). The levels of DNA methylation of Cosmc promotor region were not significantly different between the lymphocytes of the three children populations (P = 0.113), but there were significant differences between IgAN lymphocytes and lymphocytes of the other two children populations after IL-4 (P<0.0001) or AZA (P<0.0001). Cosmc mRNA expression was low in IgAN lymphocytes compared to the other two groups (P<0.0001). The level of aberrantly glycosylated IgA1 was markedly higher in IgAN group compared to the other groups (P<0.0001). After treatment with IL-4, the levels of Cosmc DNA methylation and aberrantly glycosylated IgA1 in IgAN lymphocytes were remarkably higher than the other two groups (P<0.0001) with more markedly decreased Cosmc mRNA content (P<0.0001). After treatment with AZA, the levels in IgAN lymphocytes were decreased, but was still remarkably higher than the other two groups (P<0.0001), while Cosmc mRNA content in IgAN lymphocytes were more markedly increased than the other two groups (P<0.0001). The alteration of DNA methylation by IL-4 or AZA specifically correlates in IgAN lymphocytes with alterations in Cosmc mRNA expression and with the level of aberrantly glycosylated IgA1 (r = −0.948, r = 0. 707). Our results suggested that hypermethylation of Cosmc promoter region could be a key mechanism for the reduction of Cosmc mRNA expression in IgAN lymphocytes with associated increase in aberrantly glycosylated IgA1.

Analysis of regulatory T cell subsets in the peripheral blood of immunoglobulin A nephropathy (IgAN) patients.

The aim of this study was to investigate the clinical significance of regulatory T cells (Tregs) and its subsets in immunoglobulin A nephropathy (IgAN) patients. Peripheral blood samples of 20 IgAN patients and 20 healthy individuals of similar ages were analyzed. Levels of Tregs and its subsets, namely nTregs and iTregs, were analyzed using flow cytometry. The number of Tregs in IgAN patients was significantly lower than that in the healthy controls. While significant reduction in iTregs primarily contributed to this effect (P < 0.01), nTreg levels did not significantly change (P > 0.05). The levels of serum IL-17, IL-10 and TGF-β were detected by ELISA method. The levels of IL-10 and TGF-β in IgAN patients were lower (P < 0.05), whereas those of IL-17 in the IgAN group were higher (P < 0.05) than those in the controls. In conclusion, the change in Tregs count in the peripheral blood of IgAN patients is mainly caused by the reduction in iTregs, suggesting a substantial role in the prognosis and treatment of IgAN.

Toll-like receptor 4 is involved in a protective effect of rhein on immunoglobulin A nephropathy.

Objectives:The objective was to investigate the protective effects of rhein on renal histology change and the effects of rhein on renal tissue toll-like receptor (TLR) 4, TLR9, transforming growth factor-β1 (TGF-β1) expression in immunoglobulin A nephropathy (IgAN) rats.Materials and Methods:Bovine serum albumin-lipopolysaccharide-carbon tetrachloride 4 method was used to establish IgAN model. Thirty-two male sprague dawley rats were randomly divided into the control group, IgAN model group, rhein-prevented group, and rhein-treated group. 24-h urinary protein (UP), creatinine, urea, alanine aminotransferase (ALT), total protein (TP) contents in the serum of rats were detected with automatic biochemical analyzer and renal pathological changes were observed by the hematoxylin and eosin and periodic acid-Schiff staining. The glomerular deposition of IgA was measured by immunofluorescence staining. Real-time polymerase chain reaction and immunohistochemistry were used to detect renal tissue contents of TLR4, TLR9, TGF-β1 messenger ribonucleic acid and protein expression.Results:The biochemical parameters results of IgAN model rats showed that the 24-h UP excretion and ALT concentration were much higher, and TP concentration was much lower than those of the control group (P < 0.05). Granule-like or mass-like IgA depositions in the mesangial area, glomerular hypercellularity, hyperplasia of mesangial matrix, and tubulointerstitial fibrosis were found in IgAN group. Rhein-prevented and rhein-treated both improved the biochemical parameters and relieved renal pathological injury. The expressions of renal tissue TLR4, TGF-β1, but not TLR9 were significantly elevated in IgAN model rats (P < 0.05). Rhein-prevented and rhein-treated both inhibited TLR4 and TGF-β1 expressions.Conclusion:Rhein significantly improved the serum and urine biochemical parameters, and attenuated the glomerular pathological changes and tubulointerstitial fibrosis in IgAN rats. The mechanism may involve inhibition of renal TLR4 and TGF-β1 secretion.

Relationship between rs1047763 polymorphism of the C1GALT1 gene and susceptibility to immunoglobulin A nephropathy in Xinjiang Uyghur people.

We explored the relationship between rs1047763, a single-nucleotide polymorphism (SNP) of the C1GALT1 gene, and genetic susceptibility to immunoglobulin A nephropathy (IgAN) in Xinjiang Uyghur people. The study comprised 90 patients with IgAN and 90 normal controls recruited from Uyghur people. The distribution of the rs1047763 polymorphism of C1GALT1 in each group was determined by direct sequencing analysis. The gene type, gene frequency, allele type, and allele frequency were calculated by direct counting and the genotype was investigated using the Hardy-Weinberg equilibrium test. The SPSS17.0 software was used for data processing, and genotype and allele frequencies were compared using the χ2 test. In the IgAN group, the AA, AG, and GG genotype frequencies in the rs1047763 polymorphism of the C1GALT1 gene were 21.10, 47.80, and 31.10%, respectively, while AA, AG, and GG genotype frequencies in the control group were 17.8, 40.0, and 42.2%, respectively. There was no statistically significant difference between the two groups (P > 0.05). The rs1047763 SNP of the C1GALT1 gene probably has no correlation with genetic susceptibility to IgAN in Xinjiang Uyghur people.

Distribution of Streptococcus mutans strains with collagen-binding proteins in the oral cavity of IgA nephropathy patients

IgA nephropathy (IgAN) is the most common primary chronic glomerulonephritis; however, its precise initiating pathogenesis remains unclear. Streptococcus mutans is a major pathogen of human dental caries. S. mutans strains with the cnm gene encoding Cnm, a collagen-binding protein, have been reported to contribute to the development of systemic diseases. However, the contribution of S. mutans with Cnm in the development of IgAN has not been reported. The aim of this study was to investigate the prevalence of cnm-positive S. mutans in IgAN patients and clarify the effects of cnm-positive S. mutans on the histological pathology of IgAN. We identified the cnm gene in S. mutans isolated in saliva specimens, which were collected from IgAN patients (n = 53) and control subjects (n = 50). We evaluated the collagen-binding properties of S. mutans in IgAN patients and controls. The clinical parameters and histological scores were also assessed in IgAN patients. The rates of S. mutans isolation in IgAN and control groups were 84.0 and 84.9 %, respectively, not significantly dfferent. cnm-positive strains were significantly more prevalent in the IgAN group than in controls (32.1 vs. 14.0 %, p < 0.05). With regard to collagen-binding assays, the binding rates of cnm-positive strains were significantly higher in the IgAN group than in controls (96.6 vs. 30.0, p < 0.05). In addition, the segmental glomerulosclerosis scores were significantly higher in cnm-positive patients with IgAN than in cnm-negative patients with IgAN (0.94 vs. 0.57, p < 0.05). cnm-positive S. mutans strains are potentially associated with the pathogenesis of IgAN.

Expression and role of transforming growth factor-β1 and matrix metalloproteinase 9 in IgA nephropathy children

Objective To study the association of transforming growth factor-β1(TGF-β1), matrix metalloproteinase 9(MMP-9)in serum, urinary and renal tissue of IgA nephropathy(IgAN) in children.In order to explore the noninvasive and easy repeatable biomarker, for assessing the severity and disease progression in IgAN children. Me-thods Fifty-four children with biopsy-proven IgAN and 55 healthy normal children(healthy control group) were enrolled between Jul.2009 and Jan.2013.The IgAN group was divided into 3 clinical groups according to their clinical features: isolated hematuria(IH) group(IH group, 20 cases), hematuria and proteinuria(HP) group(HP group, 16 cases), and nephritic syndrome group(NS group, 18 cases). Patients were divided into 2 groups according to their Lee's pathologic classification: grade Ⅰ+ Ⅱ group(Lee's Ⅰ+ Ⅱ, 39 cases), and grade Ⅲ+ Ⅳ group(Lee's Ⅲ+ Ⅳ, 15 cases). There were no grade Ⅴ in the study.The injury of renal tubules and interstitium was divided into 0-Ⅲ grades according to Katafuchi pathologic classification.IgAN group was also divided into 4 groups according to their immunophenotype: IgA(7 cases), IgA+ IgG group(9 cases), IgA+ IgM group(10 cases), IgA+ IgG+ IgM group(28 cases). The expressions of TGF-β1 and MMP-9 of each group were measured by enzyme-linked immunosorbent assay(ELISA). Immunohistochemistry was used to measure the expressions of TGF-β1 and MMP-9 in the paraffin sections of 34 IgAN and 3 normal renal tissue, and ELISA was used to measure the levels of TGF-β1 and MMP-9 in serum and urine.SPSS 16.0 statistical software was used. Results The levels of TGF-β1 and MMP-9 in serum and urinary of the IgAN group were significantly higher than those in the healthy control group(all P 0.05). The serum TGF-β1 and MMP-9 levels in Lee's Ⅲ+ Ⅳ group were significantly higher than those in Lee's Ⅰ+ Ⅱ group(all P 0.05). The serum and urinary TGF-β1 and MMP-9 levels were no statistically significant in all immune classification(all P>0.05). The expressions of TGF-β1 and MMP-9 in renal tubules and interstitium of Katafuchi Ⅲ grade were significantly enhanced than those in Katafuchi Ⅰ grade. Conclusions The serum TGF-β1 and MMP-9 play an important role in the severity and progression of children with IgAN.TGF-β1 and MMP-9 may be two more noninvasive and easy repeatable biomarkers to be used in evaluating the progression of IgAN in children. Key words: IgA nephropathy; Transforming growth factor-β1; Matrix metalloproteinase 9; Renal tissue; Child

Tonsillectomy and IgA Nephropathy Recurrence After Kidney Transplantation.

[Background and purpose] The pathophysiology of IgA nephropathy (IgAN) remains poorly understood, although it frequently recurs after kidney transplantation, and its recurrence has been reported to depend on some factors. We studied a retrospective single-center analysis of renal transplant recipients with IgAN as the cause of end-stage renal failure to examine factors that may affect IgAN recurrence after kidney transplantation and the preventative and therapeutic effects of tonsillectomy on IgAN recurrence. [Material and method] From January 1987 to December 2012, 126 kidney transplantations were performed at Ryuku University Hospital, among which 14 cases (10 men; and 4 women) were identified with IgAN as their primary disease. Their mean age at transplantation was 38.1 ± 8.4 years.Donor types were as follows: 2 cadaveric, and 12 living (10 related; 2 unrelated). Tonsillectomy was performed for 8 cases; 2 were for therapeutic purposes for IgAN recurrence, and 6 were to prevent IgAN recurrence. IgAN recurrence diagnosis was based on light microscopic findings showing mesangioproliferative changes and mesangial IgA deposition on the basis of immunofluorescence analysis. [Result] Among the 14 patients, 2 cases were identified as IgAN recurrence based on episode biopsies. Both the recurrent cases had living related kidney donors. One patient with recurrent transplant IgAN progressed to graft failure 2.0 years after tonsillectomy, whereas the other subsequently retained renal function for 7.0 years. No pathological IgAN recurrence was found in 6 preventative tonsillectomy cases and in 2 cadaveric transplants that did not undergo tonsillectomy. Among 6 preventative tonsillectomy patients, 5 had improved urinary findings (protein urine and/or microscopic hematuria) after tonsillectomy. The 10 years living kidney donor graft survival rate of the IgAN group was lower than that of the non IgAN group (50 % vs. 88 %; P< 0.05). [Conclusion] Our data suggest that IgAN recurs more frequently after living related kidney transplantation than with cadaveric and living unrelated donor transplantation. Furthermore, no pathological IgAN recurrence was found after preventative tonsillectomy, and it was found that tonsillectomy could improve urinary findings. Thus, tonsillectomy has preventive and/or therapeutic effects for IgAN recurrence after kidney transplantation.

Complement components, proteolysis‑related, and cell communication‑related proteins detected in urine proteomics are associated with IgA nephropathy.

IgA nephropathy (IgAN) is the most common primary glomerulonephritis. The first symptoms of IgAN are erytrocyturia or hematuria, proteinuria, and decline in renal function, or any combination of the above. One of the promising diagnostic methods is urine proteomics. We studied urine proteomics in patients with IgAN and age- and sex‑matched healthy controls. To minimize the risk of protein degradation, we proposed a new protocol for urine collection and preparation. A total of 30 patients with IgAN and 30 controls were enrolled into the study. Thirty urine samples of the IgAN group were divided into 3 disease pooled samples (DPS I, II, and III) and 30 urine samples of the control group were divided into 3 control pooled samples (CPS I, II, and III). We used isoelectric focusing/liquid chromatography-mass spectrometry/mass spectrometry (IEF/LC‑MS/MS) to detect all proteins larger than 10 kDa. Using qualitative analysis, we identified 761, 951, and 956 proteins in each of the 3 IEF/LC‑MS/MS experiments. The results were combined, yielding a dataset with 1238 proteins identified by at least 2 peptides. The statistical analysis of the quantitative results revealed 18 proteins that were differently populated in the urine of IgAN patients compared with healthy controls. We found increased urinary concentrations of complement components, coagulation factors, extracellular matrix, intracellular, transmembrane, and other proteins in patients with IgAN. Some of them have never been linked to IgAN before. We demonstrated that urine proteomics is a promising tool for diagnosing and monitoring patients with IgAN.

Expression and correlation analysis of IL-4, IFN-γ and FcαRI in tonsillar mononuclear cells in patients with IgA nephropathy

BackgroundClinical deterioration of IgA nephropathy (IgAN) is frequently preceded by episodes of upper respiratory tract infection such as tonsillitis. The aim of this study was attempt to investigate the expression and correlation of IL-4, IFN-γ and FcαRI in tonsillar mononuclear cells under stimulations of α-hemolytic streptococcus (HS) or lipopolysaccharide (LPS) in patients with IgA nephropathy. MethodsTonsillar mononuclear cells isolated from 26 patients with IgAN and 25 patients with chronic tonsillitis (CT) as controls were cultured for 72h with or without α-hemolytic streptococcus (HS) and lipopolysaccharide (LPS) stimulation. Concentration of IL-4 and IFN-γ level were determined by ELISA. Expression levels of IL-4, IFN-γ and FcαRI mRNA were measured by real-time PCR, respectively. FcαRI expressing cells were tracked by flow cytometry. ResultsThe supernatant concentration of IL-4, IFN-γ and the expression of IL-4, IFN-γ and FcαRI mRNA in the IgA nephropathy group was markedly increased compared with the non-IgAN group. With the stimulation of HS, the production of IL-4 and the FcαRI expressing cells in the IgA nephropathy group was significantly increased than the non-IgAN group, while the secretion of IFN-γ was remarkably decreased in the IgA nephropathy group than the non-IgAN group. Upon the LPS stimulation, the concentration of IL-4, IFN-γ in the supernatant of the IgA nephropathy group was markedly increased compared with the non-IgAN group. However, there wasn’t significant difference in the FcαRI expressing cells between the LPS stimulated IgAN group and the non-IgAN group. The expression of FcαRI is in a negative correlation with IL-4 while positive with IFN-γ. ConclusionThe tonsillar mononuclear cells of IgAN may in a state of overactive immune response, which probably can promote the secretion of Th cytokines, involving in the pathogenesis of IgAN.

Assessment of the effect of mesangial hypercellularity in childhood nephropathies to the clinical and laboratory findings

Aim: To assess the relationship between mesangial hypercellularity in various childhood nephropathies and clinical and laboratory parameters. Methods and patients: The reports of the renal biopsies were evaluated retrospectively. The patients with diagnosis of IgA nephropathy (isolated and Henoch–Schönlein nephritis), IgM nephropathy, or isolated mesangial proliferative glomerulonephritis were included. Each nephropathy group was divided into two subgroups according to the severity of mesangial hypercellularity as mild and severe. The biochemical data and histopathological findings of the patients were recorded. Results: When the groups were compared, it was found that the patients with IgA nephropathy had hematuria (p = 0.043) and the patients with IgM nephropathy had nephrotic syndrome more frequently than the other patients (p = 0.01). No difference was detected between the groups regarding the severity of mesangial hypercellularity. On the other hand, when the groups were evaluated within themselves, no significant association was detected between the severity of mesangial hypercellularity and clinical and laboratory parameters. It was determined that the renal biopsy was performed earlier in patients with Henoch–Schönlein nephritis compared to the other cases (p = 0.004). Compared to the isolated IgA nephropathy group, it was found that the number of cases with severe mesangial hypercellularity was higher and the level of proteinuria was more prominent in patients with Henoch–Schönlein nephritis. Additionally, when the patients with Henoch–Schönlein nephritis were evaluated, the degree of proteinuria was found to be higher in patients with severe mesangial hypercellularity compared to those of showing mild mesangial hypercellularity (p = 0.002). Conclusion: It was observed that there is no direct relation between the severity of mesangial hypercellularity and clinical and laboratory findings in various childhood nephropathies. However, when Henoch–Schönlein nephritis is compared with IgA nephropathy, it was found that the severity of mesangial hypercellularity was higher in cases with Henoch–Schönlein nephritis and the level of proteinuria was more prominent in those cases. However, no difference was detected in glomerular filtration rates and biochemical data with regard to the level of mesangial hypercellularity.

Proteomic Analysis of Whole Glomeruli in Patients with IgA Nephropathy Using Microsieving

Background: To promote understanding of immunoglobulin A nephropathy (IgAN) pathophysiology, we tried to elucidate glomerular protein profiles in IgAN, using microsieving that we established recently to isolate glomeruli from renal biopsy samples and proteomic approaches. Methods: Glomeruli were isolated from renal biopsy samples of patients with IgAN (n = 5) and with minimal change nephrotic syndrome (MCNS; n = 5) using microsieving. Proteins extracted from the isolated glomeruli were separated by 2-dimensional differential gel electrophoresis (2D-DIGE). Proteins with different amounts between the two groups were identified by mass spectrometry. One of the identified proteins, α-actinin-4 (ACTN4), was further analyzed by Western blotting, RT-polymerase chain reaction (PCR), and immunohistochemistry. Results: By 2D-DIGE, 72 out of the detected 1,170 protein spots showed significantly different intensity between the two groups (p < 0.05). Thirty-four out of the 72 protein spots showed more than 1.5-fold or less than 1/1.5-fold intensity, out of which 16 protein spots were successfully identified. No microbial protein was identified. ACTN4 molecules with a low molecular weight of approximately 77 kDa were found to increase in the IgAN group. Lack of an N-terminal part of ACTN4 was demonstrated by Western blotting. No defect of mRNA for ACTN4 was evidenced by RT-PCR. Predominant existence of ACTN4 in capillary walls of glomeruli of IgAN patients was demonstrated by immunohistochemistry in glomerular sections of patients with IgAN. Conclusion: Use of microsieving enabled us to biochemically analyze glomerular proteins in renal biopsy samples from patients with glomerular diseases. With this method, we demonstrated skewed glomerular protein profiles in IgAN.

Th1/Th2 polarization in tonsillar lymphocyte form patients with IgA nephropathy

Imbalance of Th1/Th2 pro-inflammatory cytokines plays an important role in the development and progression of IgA nephropathy (IgAN). Clinical development and exacerbation of IgAN are frequently preceded by episodes of upper respiratory tract infection, and palatine tonsils represent the predominant immunocompetent tissue of the upper respiratory tract. This study examined tonsillar lymphocytes of IgAN who suffered from tonsillitis (n = 22), and using tonsils derived from patients with chronic tonsillitis (n = 24) but without renal disease as a control. We identified a polarization toward Th2 response in tonsils of IgAN patients. TH0 cells are differentially mobilized during contact sensitization and by adjuvants such as lipopolysaccharide (LPS) that induce T-helper type 1 (Th1) responses, or α-hemolytic streptococcus (HS) that induces T-helper type 2 (Th2) responses. Th1:Th2 ratio is correlated with proteinuria and renal pathologic changes in IgAN group. Our study suggests that IgAN is associated with the change in Th1/Th2 balance in favor of Th2 lymphocytes.

Preliminary study of the acoustic radiation force impulse imaging in diagnosis and treatment of IgA nephropathy

Objective To investigate the value of acoustic radiation force impulse (ARFI) on evaluating the renal tubular atrophy and interstitial fibrosis of IgA nephropathy(IgAN).Methods 117 patients diagnosed with IgAN,and 80 normal cases were examined with conventional ultrasonography and ARFI,recording the shear wave velocities(SWV),the hemodynamics and the renal biopsy results to quantify the elastic changes of renal cortex,with the aim of assessing AFRI's advantages on early diagnosing the impair of kidney.Results The SWV value of renal cortex,renal medulla of IgAN group were smaller than that of normal group (P ＜0.05),while the collective system SWV value had no difference(P ＞0.05).According to the ROC curve,the cut-off value of SWV in renal cortex was 2.7 cm/s.The accuracy of the diagnosis was 71 ％,which was better than RI (61.9％).The SWV value of the renal cortex decreased with the different stage of 1,2,3,4,but stage 5 was larger than stage 4.The degree of renal tubular atrophy and interstitial fibrosis(T) can be classified by the SWV of renal cortex,and the SWV of T2 was larger than T1 (P ＜0.05).Conclusions By quantifying the elastic change of kidney,ARFI technology is able to distinguish the degree of renal tubular atrophy and interstitial fibrosis,diagnose renal impairment early and judge the clinical stages.Furthermore the sensitivity,specificity,accuracy of it are superior to hemodynamic. Key words: Ultrasonopgraphy; Glomerulonephritis, IgA; Acoustic radiation force impulse