The level of galactose-deficient IgA1 in the sera of patients with IgA nephropathy is associated with disease progression
Authors: Na Zhao, Ping Hou, Jicheng Lv, Zina Moldoveanu, Yifu Li, Krzysztof Kiryluk, Ali G. Gharavi, Jan Novak, Hong Zhang
Journal: Kidney International (2012)

Although high serum levels of galactose-deficient IgA1 (an important biomarker of IgA nephropathy (IgAN)) are found in most patients with IgAN, their relationship to disease severity and progression remains unclear. To help clarify this we prospectively enrolled 275 patients with IgAN and followed them for a median of 47 months (range 12–96 months). Serum galactose–deficient IgA1 was measured at the time of diagnosis using a lectin-based ELISA, and renal survival was modeled using the Cox proportional hazards method. The serum levels of galactose-deficient IgA1 were higher in patients with IgAN compared to those in healthy controls. Importantly, in adjusted analysis, higher levels of galactose-deficient IgA1 were independently associated with a greater risk of deterioration in renal function with a hazard ratio of 1.44 per standard deviation of the natural log–transformed galactose-deficient IgA1 concentration. In reference to the first quartile, the risk of kidney failure increased such that the hazard ratio for the second quartile was 2.47, 3.86 for the third, and 4.76 for the fourth quartile of the galactose-deficient IgA1 concentration. Hence, elevated serum levels of galactose-deficient IgA1 are associated with a poor prognosis in IgAN. Although high serum levels of galactose-deficient IgA1 (an important biomarker of IgA nephropathy (IgAN)) are found in most patients with IgAN, their relationship to disease severity and progression remains unclear. To help clarify this we prospectively enrolled 275 patients with IgAN and followed them for a median of 47 months (range 12–96 months). Serum galactose–deficient IgA1 was measured at the time of diagnosis using a lectin-based ELISA, and renal survival was modeled using the Cox proportional hazards method. The serum levels of galactose-deficient IgA1 were higher in patients with IgAN compared to those in healthy controls. Importantly, in adjusted analysis, higher levels of galactose-deficient IgA1 were independently associated with a greater risk of deterioration in renal function with a hazard ratio of 1.44 per standard deviation of the natural log–transformed galactose-deficient IgA1 concentration. In reference to the first quartile, the risk of kidney failure increased such that the hazard ratio for the second quartile was 2.47, 3.86 for the third, and 4.76 for the fourth quartile of the galactose-deficient IgA1 concentration. Hence, elevated serum levels of galactose-deficient IgA1 are associated with a poor prognosis in IgAN. IgA nephropathy (IgAN) is the most common form of primary glomerulonephritis worldwide1.Levy M. Berger J. Worldwide perspective of IgA nephropathy.Am J Kidney Dis. 1988; 12: 340-347Abstract Full Text PDF PubMed Scopus (214) Google Scholar and the leading cause of end-stage renal disease (ESRD) in young adults.2.Nair R. Walker P.D. Is IgA nephropathy the commonest primary glomerulopathy among young adults in the USA?.Kidney Int. 2006; 69: 1455-1458Abstract Full Text Full Text PDF PubMed Scopus (138) Google Scholar The pattern of glomerular immunoglobulin deposits is suggestive of an immune complex–mediated mechanism.3.Donadio J.V. Grande J.P. IgA nephropathy.N Engl J Med. 2002; 347: 738-748Crossref PubMed Scopus (681) Google Scholar, 4.Smith A.C. Molyneux K. Feehally J. et al.O-glycosylation of serum IgA1 antibodies against mucosal and systemic antigens in IgA nephropathy.J Am Soc Nephrol. 2006; 17: 3520-3528Crossref PubMed Scopus (100) Google Scholar, 5.Suzuki H. Fan R. Zhang Z. et al.Aberrantly glycosylated IgA1 in IgA nephropathy patients is recognized by IgG antibodies with restricted heterogeneity.J Clin Invest. 2009; 119: 1668-1677PubMed Google Scholar, 6.Tomana M. Matousovic K. Julian B.A. et al.Galactose-deficient IgA1 in sera of IgA nephropathy patients is present in complexes with IgG.Kidney Int. 1997; 52: 509-516Abstract Full Text PDF PubMed Scopus (280) Google Scholar, 7.Novak J. Julian B.A. Tomana M. et al.IgA glycosylation and IgA immune complexes in the pathogenesis of IgA nephropathy.Semin Nephrol. 2008; 28: 78-87Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar Multiple studies have established the contribution of aberrantly glycosylated IgA1 in the pathogenesis of IgAN.8.Amore A. Cirina P. Conti G. et al.Glycosylation of circulating IgA in patients with IgA nephropathy modulates proliferation and apoptosis of mesangial cells.J Am Soc Nephrol. 2001; 12: 1862-1871PubMed Google Scholar, 9.Barratt J. Feehally J. Smith A.C. Pathogenesis of IgA nephropathy.Semin Nephrol. 2004; 24: 197-217Abstract Full Text Full Text PDF PubMed Scopus (118) Google Scholar, 10.Barratt J. Feehally J. IgA nephropathy.J Am Soc Nephrol. 2005; 16: 2088-2097Crossref PubMed Scopus (401) Google Scholar, 11.Smith A.C. de Wolff J.F. Molyneux K. et al.O-glycosylation of serum IgD in IgA nephropathy.J Am Soc Nephrol. 2006; 17: 1192-1199Crossref PubMed Scopus (73) Google Scholar, 12.Suzuki H. Moldoveanu Z. Hall S. et al.IgA1-secreting cell lines from patients with IgA nephropathy produce aberrantly glycosylated IgA1.J Clin Invest. 2008; 118: 629-639PubMed Google Scholar IgA1 is one of the very few serum proteins with O-glycosylation. The O-glycans in the hinge region of normal IgA1 consist of N-acetylgalactosamine, galactose, and sialic acid. In patients with IgAN, some of the O-glycans of circulating IgA1 are deficient in galactose. Similarly, mesangial immune deposits eluted directly from glomeruli of IgAN patients contained galactose-deficient IgA1 (Gd-IgA1).7.Novak J. Julian B.A. Tomana M. et al.IgA glycosylation and IgA immune complexes in the pathogenesis of IgA nephropathy.Semin Nephrol. 2008; 28: 78-87Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar, 13.Allen A.C. Bailey E.M. Brenchley P.E. et al.Mesangial IgA1 in IgA nephropathy exhibits aberrant O-glycosylation: observations in three patients.Kidney Int. 2001; 60: 969-973Abstract Full Text Full Text PDF PubMed Scopus (268) Google Scholar, 14.Hiki Y. Odani H. Takahashi M. et al.Mass spectrometry proves under-O-glycosylation of glomerular IgA1 in IgA nephropathy.Kidney Int. 2001; 59: 1077-1085Abstract Full Text Full Text PDF PubMed Scopus (299) Google Scholar On the basis of the potential pathogenic role of Gd-IgA1 in the development of IgAN, a quantitative assay for serum Gd-IgA1 holds promise as a non-invasive diagnostic tool. We previously reported an increased binding of a N-acetylgalactosamine-specific lectin from Helix aspersa (HAA) to desialylated serum Gd-IgA1 in Caucasian patients with IgAN.15.Moldoveanu Z. Wyatt R.J. Lee J.Y. et al.Patients with IgA nephropathy have increased serum galactose-deficient IgA1 levels.Kidney Int. 2007; 71: 1148-1154Abstract Full Text Full Text PDF PubMed Scopus (292) Google Scholar Our earlier study demonstrated that Gd-IgA1 was also closely associated with the pathologic phenotype of IgAN.16.Xu L.X. Zhao M.H. Aberrantly glycosylated serum IgA1 are closely associated with pathologic phenotypes of IgA nephropathy.Kidney Int. 2005; 68: 167-172Abstract Full Text Full Text PDF PubMed Scopus (88) Google Scholar However, it is not known whether elevated levels of Gd-IgA1 in sera of IgAN patients are associated with accelerated disease progression, or a worse prognosis.17.Coppo R. Amore A. Aberrant glycosylation in IgA nephropathy (IgAN).Kidney Int. 2004; 65: 1544-1547Abstract Full Text Full Text PDF PubMed Scopus (152) Google Scholar We have previously established a large prospective cohort of patients with IgAN followed regularly at the Peking University Institute of Nephrology. This cohort has been assembled and followed as part of the Peking University IgAN database project since 2003 (http://www.renal-online.org), with blood and urine samples collected at the time of kidney biopsy, and clinical data collected prospectively for all enrolled patients. In this study, we utilize the above resource to examine the prognostic utility of serum levels Gd-IgA1. Among the 275 IgAN patients, there were 147 (53.5%) males and 128 (46.5%) females with mean age at the time of kidney biopsy of 32.7±10.7 years. On biopsy, average proteinuria level was 1.92±1.89g per 24h (range 0.01–13.72g/24h) and average estimated glomerular filtration rate (eGFR) was 82.65±27.44ml/min per 1.73m2 (range 6.3–164.9ml/min per 1.73m2). Systolic blood pressure was 124±16mmHg, diastolic blood pressure 79±12mmHg, with 131 patients (47.6%) being hypertensive at baseline. The distribution by Haas grade I, II, III, IV, and V was 10.9%, 0.4%, 32.4%, 42.9%, and 13.5%, respectively. The median follow-up time was 47 months (range 12–96 months; Table 1). During the follow-up period, 266 (96.7%) patients received ACE inhibitors or ARBs therapy, 127 (46.2%) received oral corticosteroids alone or combined with other immunosuppressive agents. In total, 42 patients reached the composite end point of 50% decline in eGFR (n=39), ESRD (n=3), or death (n=2; both had a 50% decline in eGFR before death).Table 1Baseline clinical and laboratory data and levels of serum Gd-IgA1 in 275 patients with IgANCharacteristicsMean±s.d. (range or percentage)Age (year)32.7±10.7 (15–76)Gender (male)147/275 (53.5%)SBP (mmHg)124±16DBP (mmHg)79±12Hypertension (%)aHypertension was defined as SBP of greater than 140mmHg, and/or DBP of greater than 90mmHg at resting, or use of anti-hypertension medication.131/275 (47.6%)Initial proteinuria (g/day)1.92±1.89 <0.3 (%)15/275 (5.5%) 0.3–0.99 (%)84/275 (30.5%) 1.0–2.99 (%)130/275 (47.3%) ≥3.0 (%)46/275 (16.7%)eGFR (ml/min per 1.73m2)82.65±27.44Stages 1, 2, 3, and 4 CKD (KDOQI)bCKD stage 1, 2, 3, and 4 were divided by eGFR≥90, 60–89, 30–59, and 15–29, respectively.125 (45.7%), 92 (33.3%), 50 (18.0%), 8 (3%)Follow-up interval (month, median, IQR)47 (30–60)Gd-IgA1 (U/ml, median, IQR)312.5 (236.5–407.8)Total IgA (μg/ml, median, IQR)2630.0 (2125.0–3510.0)Plasma IgA1 (μg/ml, median, IQR)1709.9 (1376.2–2159.7)Therapy (%) ACE inhibitors or ARBs96.7 Prednisone46.2 Any other immunosuppressive agents (cyclophosphamide, MMF, or others)29.8Histological grading (%)cHistological grading was classified according to pathological scheme proposed by Haas.45 I, II, III, IV, V30 (10.9%), 1 (0.36%), 89 (32.4%), 118 (42.9%), 37 (13.5%)Abbreviations: ACE, angiotensin-converting enzyme; ARB, angiotensin II receptor blocker; CKD, chronic kidney disease; DBP, diastolic blood pressure; eGFR, estimate glomerular filtration rate; Gd-IgA1, galactose-deficient IgA1; IgAN, IgA nephropathy; IQR, interquartile range; MMF, mycophenolate mofetil; SBP, systolic blood pressure.a Hypertension was defined as SBP of greater than 140mmHg, and/or DBP of greater than 90mmHg at resting, or use of anti-hypertension medication.b CKD stage 1, 2, 3, and 4 were divided by eGFR≥90, 60–89, 30–59, and 15–29, respectively.c Histological grading was classified according to pathological scheme proposed by Haas.45.Haas M. Histologic subclassification of IgA nephropathy: a clinicopathologic study of 244 cases.Am J Kidney Dis. 1997; 29: 829-842Abstract Full Text PDF PubMed Scopus (399) Google Scholar Open table in a new tab Abbreviations: ACE, angiotensin-converting enzyme; ARB, angiotensin II receptor blocker; CKD, chronic kidney disease; DBP, diastolic blood pressure; eGFR, estimate glomerular filtration rate; Gd-IgA1, galactose-deficient IgA1; IgAN, IgA nephropathy; IQR, interquartile range; MMF, mycophenolate mofetil; SBP, systolic blood pressure. HAA-based enzyme-linked immunosorbent assay (ELISA) for Gd-IgA1 was highly reproducible (r=0.903, P<0.0001). The level of Gd-IgA1 in IgAN patients (median: 312.5U/ml, interquartile range (IQR) 236.5–407.8U/ml) was significantly higher compared with that of the healthy controls (148.3U/ml, IQR 106.1–209.1U/ml, P<0.001). The patients were next divided into four equal groups according to the quartiles of the Gd-IgA1 distribution; group 1: serum Gd-IgA1 levels <236.5U/ml; group 2: serum Gd-IgA1 between 236.5 and 312.5U/ml; group 3: serum Gd-IgA1 between 312.5 and 407.8U/ml; and Group 4: serum Gd-IgA1 >407.8U/ml. Among cases, we observed no significant association of Gd-IgA1 levels with age, gender, or body mass index (BMI; Table 2). There was a trend for lower eGFR, higher proteinuria, and increased use of immunosuppressives in groups 3 and 4 compared with groups 1 or 2, albeit none of these differences were statistically significant. There were also no significant differences between the groups in the blood pressure control (P=0.96) or proteinuria during follow-up (P=0.94).Table 2Baseline data of clinical features and levels of serum Gd-IgA1 of four groups defined by quartiles of serum Gd-IgA11234U/ml, median (IQR)U/ml, median (IQR)U/ml, median (IQR)U/ml, median (IQR)Group193.9 (158.4, 213.8)272.5 (247.2, 289.0)345.7 (324.2, 369.9)487.4 (438.8, 540.3)P valueNumber69696869Gender (male)41/6937/6933/6836/690.638Age (year)32.78±11.4333.87±10.6332.47±11.331.80±9.340.718Hypertension (%)34 (49.3%)34 (49.3%)27 (39.7%)36 (52.2%)0.488Proteinuria (%, >1g/day)42/69 (60.9%)39/69 (56.5%)50/68 (73.5%)45/69 (65.2%)0.196eGFR (ml/min per 1.73m2), stages 1, 2, 3, and 4 CKD (KDOQI)29/28/10/234/18/16/129/24/13/230/22/13/40.715Histological grading (mild lesions/severe lesions)26/4329/4034/3431/380.525Prednisone27 (39.13%)29 (42.03%)37 (54.41%)34 (49.28%)0.264Any other immunosuppressive agents21 (30.4%)18 (26.1%)16 (23.5%)24 (34.7%)0.481ACE inhibitors or ARBs68 (98.60%)69 (100.0%)66 (97.10%)64 (94.10%)0.157Total IgA (μg/ml, median, IQR)2026 (1566.5,2411)2479 (2309.5,2849)2880 (2362.5,3669)3483 (2855,4012)<0.001Plasma IgA1 (μg/ml, median, IQR)1352.2 (1138.3,1699.7)1574.7 (1325.9,1827.6)1868.8 (1556.8,2289.6)2109.5 (1765.5,2558.6)<0.001Kidney failure events (%)6 (8.69%)6/69 (8.69%)15/68 (22.1%)15/69 (21.7%)0.026Abbreviations: ACE, angiotensin-converting enzyme; ARB, angiotensin II receptor blocker; CKD, chronic kidney disease; eGFR, estimate glomerular filtration rate; Gd-IgA1, galactose-deficient IgA1; IQR, interquartile range.Histological grading was divided into mild lesions and severe lesions group. The former group included Haas I, II, and III, and the latter group included IV and V. Continuous data were compared by one-way analysis of variance test; dichotomous or categorical data were analyzed by χ2 test. Open table in a new tab Abbreviations: ACE, angiotensin-converting enzyme; ARB, angiotensin II receptor blocker; CKD, chronic kidney disease; eGFR, estimate glomerular filtration rate; Gd-IgA1, galactose-deficient IgA1; IQR, interquartile range. Histological grading was divided into mild lesions and severe lesions group. The former group included Haas I, II, and III, and the latter group included IV and V. Continuous data were compared by one-way analysis of variance test; dichotomous or categorical data were analyzed by χ2 test. Baseline clinical and pathological variables were tested for association with the primary progression outcome using Cox proportional hazards model. In univariate analyses, lower baseline eGFR, higher histological grading, greater degree of proteinuria, use of steroids, and higher levels of Gd-IgA1 were all significantly associated with a progression risk, but age, gender, hypertension, hemoglobin, or albumin levels were not (Supplementary Table S1 online). Download .doc (.04 MB) Help with doc files Supplementary Table S1 Unadjusted and multivariable adjusted hazard ratios (HRs) for composite of end point are presented in Table 3 according to baseline Gd-IgA1, expressed as a continuous variable or in quartiles. In adjusted analysis including baseline proteinuria, hypertension, eGFR, Haas classification, and steroid therapy, higher levels of Gd-IgA1 were independently associated with a greater risk of kidney failure (HR, per s.d. of natural log–transformed Gd-IgA1 1.44; 95% confidence interval (CI), 1.11–1.88, P=0.006). Gd-IgA1 level remained strongly associated with progression even after adjustment for time-average proteinuria and other baseline risk factors including hypertension and eGFR (HR=1.39, 95% CI 1.24–1.56, P=1.15 × 10-8). Compared with the first quartile of Gd-IgA1 (reference), the risk of kidney failure increased by quartile of Gd-IgA1 level: the HRs were 2.47 (95% CI, 0.91–6.72) for the second quartile, 3.86 (95% CI, 1.33–11.33) third quartile, and 4.76 (95% CI, 1.61–14.09) fourth quartile.Table 3Risks of composite end-point natural log–transformed Gd-IgA1 and ascending quartilesGd-IgA1, median (range), U/mlUnadjustedHazard ratio (95% confidence interval) and P-valueModel 1aModel 1 adjusted for estimate glomerular filtration rate (eGFR), proteinuria, and hypertension (yes or no). Hypertension (yes or no) was analyzed as dichotomous data.Model 2bModel 2 adjusted for covariates in model 1 plus histological grading (mild and severe lesion group). The latter variable was analyzed as categorical data.Model 3cModel 3 adjusted for covariates in model 2 plus steroid use (yes or no). The latter variable was analyzed as dichotomous data. Composite end point was defined as 50% decline of eGFR (n=29), end-stage renal disease (n=3), or death (n=2). The two deaths also had 50% decline of eGFR. Unadjusted model analyzed Gd-IgA1 as continuous data.Composite end point Per 1s.d. lnGd-IgA1312.5 (89.0–1442.0)2.07 (1.53–2.78)1.51 (1.16–1.97)1.50 (1.15–1.96)1.44 (1.11–1.88)1.68 × 10-60.0020.0030.006Gd-IgA1 quartiles 1193.88 (89.0–237.0)1 (Reference)1 (Reference)1 (Reference)1 (Reference) 2272.51 (239.0–312.0)2.63 (0.94–7.36)2.71 (0.99–7.39)2.73 (0.99–7.45)2.47 (0.91–6.72)0.0660.0520.0510.077 3345.67 (313.0–406.0)4.03(1.36–11.96)3.74 (1.28–10.93)3.72 (1.27–10.89)3.86 (1.33–11.33)0.0120.0160.0160.013 4487.36 (408.0–1442.0)6.76 (2.23–20.19)5.18 (1.75–15.34)5.29 (1.78–15.73)4.76 (1.61–14.09)0.0010.0030.0030.005Abbreviations: Gd-IgA1, galactose-deficient IgA1; lnGd-IgA1, natural log–transformed Gd-IgA1.a Model 1 adjusted for estimate glomerular filtration rate (eGFR), proteinuria, and hypertension (yes or no). Hypertension (yes or no) was analyzed as dichotomous data.b Model 2 adjusted for covariates in model 1 plus histological grading (mild and severe lesion group). The latter variable was analyzed as categorical data.c Model 3 adjusted for covariates in model 2 plus steroid use (yes or no). The latter variable was analyzed as dichotomous data.Composite end point was defined as 50% decline of eGFR (n=29), end-stage renal disease (n=3), or death (n=2). The two deaths also had 50% decline of eGFR. Unadjusted model analyzed Gd-IgA1 as continuous data. Open table in a new tab Abbreviations: Gd-IgA1, galactose-deficient IgA1; lnGd-IgA1, natural log–transformed Gd-IgA1. As shown in Figure 1, the renal survival deteriorated by the quartile of serum Gd-IgA1 level. The renal survival at first and third year in each group of patients was 100.0% and 96.9%; 100.0% and 91.8%; 100.0% and 92.2%; 98.6% and 88.6%, respectively (log-rank test, P=0.004). We also analyzed Gd-IgA1/IgA1 ratio and Gd-IgA1/total IgA ratio instead of Gd-IgA1. Gd-IgA1/IgA1 and Gd-IgA1/IgA ratios also represented risk factors for kidney disease progression in the univariate analysis (unadjusted HR=1.42, 95% CI 1.05–1.92, P=0.021; and HR=1.56, 95% CI 1.15–2.12, P=0.005, respectively) but were not significantly associated after adjustment for other covariates in the multivariate model (P=0.2 and 0.34, respectively). IgAN can have a highly variable course. Some patients progress rapidly to ESRD, whereas others have a rather benign disease trajectory.17.Coppo R. Amore A. Aberrant glycosylation in IgA nephropathy (IgAN).Kidney Int. 2004; 65: 1544-1547Abstract Full Text Full Text PDF PubMed Scopus (152) Google Scholar,18.Novak J. Tomana M. Kilian M. et al.Heterogeneity of O-glycosylation in the hinge region of human IgA1.Mol Immunol. 2000; 37: 1047-1056Crossref PubMed Scopus (59) Google Scholar The individual outcomes of IgAN remain difficult to predict. Previous longitudinal studies have associated proteinuria, hypertension, and impaired renal function at the time of diagnosis with IgAN progression.19.Bartosik L.P. Lajoie G. Sugar L. et al.Predicting progression in IgA nephropathy.Am J Kidney Dis. 2001; 38: 728-735Abstract Full Text Full Text PDF PubMed Scopus (309) Google Scholar In this study, we hypothesize that the increased serum level of Gd-IgA1, the key pathogenic feature in IgAN, causes accelerated disease progression in IgAN. On the basis of the comparison of Gd-IgA1 levels between the IgAN patients and a large group of healthy population controls, our study confirmed that serum Gd-IgA1 levels are elevated in IgAN. Most importantly, our study also demonstrated that high levels of Gd-IgA1 were predictive of renal function decline in IgAN. On the basis of our prospective follow-up data, we found that elevated level of serum Gd-IgA1 was independently associated with increased risk of renal disease progression. Among cases with biopsy-based diagnosis of IgAN, participants with the highest vs. the lowest quartile demonstrated a 4.76-fold greater risk of kidney progression. Thus, our data suggest that high levels of Gd-IgA1 may have prognostic utility in IgAN. Our observations that individuals with higher levels of Gd-IgA1 are more prone to disease progression may justify closer follow-up and possibly more aggressive treatment of such individuals. A recent study from Japan found that Gd-IgA1 was also significantly elevated in Japanese IgAN patients,20.Shimozato S. Hiki Y. Odani H. et al.Serum under-galactosylated IgA1 is increased in Japanese patients with IgA nephropathy.Nephrol Dial Transplant. 2008; 23: 1931-1939Crossref PubMed Scopus (54) Google Scholar however, this study did not examine its impact on kidney disease progression. Another study investigated the impact of Gd-IgA1 levels on the renal function decline rate among 62 Caucasian patients with IgAN.21.Camilla R. Suzuki H. Dapra V. et al.Oxidative stress and galactose-deficient IgA1 as markers of progression in IgA nephropathy.Clin J Am Soc Nephrol. 2011; 6: 1903-1911Crossref PubMed Scopus (85) Google Scholar Gd-IgA1 was independently associated with proteinuria level during the follow-up. Although Gd-IgA1 by itself was not associated with the rate of GFR decline, the combination of a high level of Gd-IgA1 and advanced oxidation protein products contributed to a more rapid loss of renal function. However, the small sample size was the major limitation of the prior studies. Taken together, these studies suggest that aberrant glycosylation of IgA1 contributes to the risk of nephropathy in both Chinese and Caucasian populations. Both in vivo and in vitro studies have now firmly established that altered O-glycosylation of serum IgA1 has a central role in the development of IgAN.22.Barratt J. Smith A.C. Feehally J. The pathogenic role of IgA1 O-linked glycosylation in the pathogenesis of IgA nephropathy.Nephrology. 2007; 12: 275-284Crossref PubMed Scopus (51) Google Scholar, 23.Tam K.Y. Leung J.C. Chan L.Y. et al.Macromolecular IgA1 taken from patients with familial IgA nephropathy or their asymptomatic relatives have higher reactivity to mesangial cells in vitro.Kidney Int. 2009; 75: 1330-1339Abstract Full Text Full Text PDF PubMed Scopus (32) Google Scholar, 24.Novak J. Tomana M. Matousovic K. et al.IgA1-containing immune complexes in IgA nephropathy differentially affect proliferation of mesangial cells.Kidney Int. 2005; 67: 504-513Abstract Full Text Full Text PDF PubMed Scopus (173) Google Scholar Novak et al. have demonstrated that B-cell abnormal expression/activity of enzymes involved in sialylation and/or galactosylation of the IgA1 hinge region O-linked glycans underlie this defect. Furthermore, anti-glycan antibodies (IgG and/or IgA1) recognize Gd-IgA1 (auto-antigen) and trigger the formation of IgA1-containing immune complexes.5.Suzuki H. Fan R. Zhang Z. et al.Aberrantly glycosylated IgA1 in IgA nephropathy patients is recognized by IgG antibodies with restricted heterogeneity.J Clin Invest. 2009; 119: 1668-1677PubMed Google Scholar In addition, it appears that the defect in the glycosylation of IgA1 is, to a large degree, genetically determined.25.Gharavi A.G. Moldoveanu Z. Wyatt R.J. et al.Aberrant IgA1 glycosylation is inherited in familial and sporadic IgA nephropathy.J Am Soc Nephrol. 2008; 19: 1008-1014Crossref PubMed Scopus (189) Google Scholar, 26.Kiryluk K. Moldoveanu Z. Sanders J.T. et al.Aberrant glycosylation of IgA1 is inherited in both pediatric IgA nephropathy and Henoch-Schonlein purpura nephritis.Kidney Int. 2011; 80: 79-87Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar, 27.Lin X. Ding J. Zhu L. et al.Aberrant galactosylation of IgA1 is involved in the genetic susceptibility of Chinese patients with IgA nephropathy.Nephrol Dial Transplant. 2009; 24: 3372-3375Crossref PubMed Scopus (54) Google Scholar However, this IgA1 O-glycosylation aberrancy (Hit 1) itself is not sufficient to induce renal injury.25.Gharavi A.G. Moldoveanu Z. Wyatt R.J. et al.Aberrant IgA1 glycosylation is inherited in familial and sporadic IgA nephropathy.J Am Soc Nephrol. 2008; 19: 1008-1014Crossref PubMed Scopus (189) Google Scholar Synthesis and binding of antibodies directed against Gd-IgA1 are required for the formation of immune complexes that accumulate in the glomerular mesangium (Hits 2).28.Suzuki H. Kiryluk K. Novak J. et al.The pathophysiology of IgA nephropathy.J Am Soc Nephrol. 2011; 22: 1795-1803Crossref PubMed Scopus (465) Google Scholar On the basis of the results of this study, we speculate that a higher level of circulating Gd-IgA1, when exposed to the second hit, may result in more severe kidney injury. The limitations of our study include the relatively short follow-up, with a median of ∼4 years. This reduces the study power, with fewer end points observed during this time period. We used a composite end point of 50% decline from baseline eGFR, ESRD, or death, which is a robust outcome used in many clinical trials.29.Manno C. Torres D.D. Rossini M. et al.Randomized controlled clinical trial of corticosteroids plus ACE-inhibitors with long-term follow-up in proteinuric IgA nephropathy.Nephrol Dial Transplant. 2009; 24: 3694-3701Crossref PubMed Scopus (203) Google Scholar Future extension of this study to a follow-up period that exceeds 10 years would enable examination of harder end points (such as ESRD alone) and provide more accurate effect estimates. Other limitations stem from the complicated multistep ELISA assay used in this study to quantify Gd-IgA1 levels. This method may not be ideal for accurate detection of small differences in Gd-IgA1 levels. Mass spectrometry–based methods, such as those30.Wada Y. Dell A. Haslam S.M. et al.Comparison of methods for profiling O-glycosylation: Human Proteome Organisation Human Disease Glycomics/Proteome Initiative multi-institutional study of IgA1.Mol Cell Proteomics. 2010; 9: 719-727Crossref PubMed Scopus (129) Google Scholar, 31.Takahashi K. Wall S.B. 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Finally, we recognize that our results may not be applicable to all patient populations, as our study cohort consists of only Han Chinese patients followed at a single tertiary care medical center. Additional longitudinal studies of more diverse patient cohorts are needed to validate our findings. Patients with IgAN are characterized by a highly variable clinical course ranging from a totally benign incidental condition to rapidly progressive renal failure. Many studies have focus on predicting outcomes in IgAN from the baseline clinical or pathological variables.19.Bartosik L.P. Lajoie G. Sugar L. et al.Predicting progression in IgA nephropathy.Am J Kidney Dis. 2001; 38: 728-735Abstract Full Text Full Text PDF PubMed Scopus (309) Google Scholar, 20.Shimozato S. Hiki Y. Odani H. et al.Serum under-galactosylated IgA1 is increased in Japanese patients with IgA nephropathy.Nephrol Dial Transplant. 2008; 23: 1931-1939Crossref PubMed Scopus (54) Google Scholar, 34.Berthoux F. Mohey H. 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