Immunofluorescence-based reclassification of idiopathic membranoproliferative glomerulonephritis: Insights from a single-center cohort

Abstract

BACKGROUND

Membranoproliferative glomerulonephritis (MPGN) is a rare form of glomerular injury with variable prognosis, defined by immunoglobulin and/or complement deposits in the mesangium and basement membranes on immunofluorescence microscopy.

AIM

To categorize it into immune complex-mediated MPGN (IC-MPGN) or complement-mediated MPGN (C-MPGN) types and to evaluate the clinical significance of this classification.

METHODS

This retrospective cohort study evaluated biopsy-confirmed primary MPGN cases in adults diagnosed between January 2010 and December 2021 at the Department of Nephrology, Sindh Institute of Urology and Transplantation, Karachi, Pakistan. Patients were followed for at least 36 months post-biopsy, and secondary causes were excluded. Based on immunofluorescence microscopy findings, cases were categorized as IC-MPGN or C-MPGN. Clinicopathological features and renal outcomes were compared between the two groups.

RESULTS

A total of 128 patients with primary MPGN were included, of whom 100 (78.1%) were reclassified as IC-MPGN and 28 (21.9%) as C-MPGN. No statistically significant differences were observed between the groups in gender distribution, clinical presentation, biopsy indications, histopathological findings, or renal function at diagnosis, except for a modest age difference (median 33.5 years vs 38 years, P = 0.029). Vascular pathology was significantly more frequent in C-MPGN, and C3 deposition was stronger and serum C3 levels lower, consistent with complement consumption. Immunosuppressive therapy was administered to 51 (51%) IC-MPGN patients and 19 (67.8%) C-MPGN patients. At 36 months, IC-MPGN patients showed numerically higher rates of complete or partial remission (62% vs 60.7%), though this difference did not reach statistical significance (log-rank test: P = 0.40). Progression to end-stage kidney disease and dialysis dependence was comparable between the two groups.

CONCLUSION

The majority of MPGN cases were reclassified as IC-MPGN, which showed numerically higher remission rates. Overall, the clinicopathological features at diagnosis were similar between the two groups, though C-MPGN, a less frequent subtype, was associated with vascular pathology and stronger complement deposition, and a poorer medium-term prognosis.

Key Words: End-stage kidney disease; Membranoproliferative glomerulonephritis; Immune complex-mediated membranoproliferative glomerulonephritis; Complement-mediated membranoproliferative glomerulonephritis; Immunofluorescence

Core Tip: Careful subclassification of membranoproliferative glomerulonephritis (MPGN) using immunofluorescence is essential, as immune complex-mediated MPGN and complement-mediated MPGN (C-MPGN) types differ in underlying mechanisms and outcomes. Although clinical and histopathological features at presentation may appear similar, subtle distinctions, such as stronger C3 deposition and lower serum C3 in C-MPGN, may provide a clue for accurate diagnosis. Immune complex-mediated MPGN is more common and tends to achieve higher remission rates, while C-MPGN may carry a poorer prognosis despite comparable progression to end-stage kidney disease. Long-term follow-up and tailored immunosuppressive strategies are critical for both subtypes to optimize renal outcomes and better understand disease trajectories in diverse populations.

INTRODUCTION

Membranoproliferative glomerulonephritis (MPGN) is a histological pattern of glomerular injury characterized by mesangial hypercellularity and endocapillary proliferation, with lobular prominence, double contours, and a tram-track appearance visible on periodic acid-Schiff and silver stains[1-3].

Historically, MPGN has been categorized into three types based on the location of immune deposits observed on electron microscopy. These include MPGN type I, type II, and type III, recognized by the presence of electron-dense deposits in the subendothelial, intramembranous (within the lamina densa), or both subendothelial and subepithelial locations, respectively[4-6]. However, this morphological classification provides little insight into the underlying pathophysiology, which is crucial for therapeutic strategies, prognosis, and understanding the etiopathogenesis of the disease to guide proper patient management.

Sethi and Fervenza[4] proposed an etiology- and pathophysiology-based classification of MPGN, categorizing lesions into immune complex-mediated MPGN (IC-MPGN) and complement-mediated MPGN (C-MPGN), or, more broadly, C3 glomerulopathy (C3G), based on immunofluorescence (IF) findings[5]. This classification highlights distinctive pathogenetic pathways underlying the two disease processes.

Immune complex-mediated injury, the more common form, results from activation of the classical complement pathway due to persistent antigenemia and antigen-antibody complex formation, accompanied by substantial immunoglobulin deposition. In adults, the common causes are infections, autoimmune diseases, or monoclonal gammopathies, whereas in children, the etiology is often idiopathic.

In contrast, C3G results from disordered regulation of the alternative complement pathway[1]. IF demonstrates exclusive or predominant C3 staining, often two orders of magnitude stronger than any other immune reactant (scored on a scale of 0-3+), with minimal or nil immunoglobulin deposition[5]. The primary cause of C3G is the presence in the circulation of autoantibodies against convertase enzymes (C3, C4, or C5 nephritic factors), which prevent normal degradation of the alternative pathway C3 convertase. Other causes include genetic mutations affecting complement regulatory proteins, leading to compromised function, while some cases remain idiopathic. The clinical application of this new classification allows more relevant categorization of cases without requiring electron microscopy, particularly in resource-limited centers[7].

C3G is further subdivided into dense deposit disease (DDD) and C3 glomerulonephritis (C3GN) based on an electron microscopy study[5]. Deposits within the lamina densa indicate DDD, whereas deposits in the mesangium, subendothelial, subepithelial, and/or intramembranous locations define C3GN[8].

The prevalence of MPGN has diminished in developed countries, largely due to early detection and effective treatment of infections. On the other hand, MPGN remains a substantial contributor to end-stage kidney disease (ESKD) in developing nations, including Pakistan. MPGN continues to account for approximately 7%-10% of all biopsies performed for acute glomerulonephritis in developing countries[9]. Nevertheless, data on the application of this new classification from these regions, particularly Pakistan, remain limited.

This IF-based classification carries significant prognostic and therapeutic implications. Patients with IC-MPGN often require evaluation for secondary causes, with treatment directed at addressing the underlying etiology when identified[10]. Conversely, C3G frequently follows a more aggressive course, with up to 50% of patients progressing to ESKD within 10 years[11]. Therapeutic strategies for C3G are evolving, ranging from immunosuppression to emerging complement inhibitors targeting C5, C3, factor B, or other components of the alternative pathway[12].

The aim of this study is to reclassify previously diagnosed cases of primary MPGN and to investigate the prevalence of IC-MPGN and C-MPGN in our population. As IF facilities are available in our laboratory, we are in a better position to reclassify diagnosed cases of primary MPGN according to the pathophysiology-based classification, which may assist clinicians in selecting proper management strategies.

MATERIALS AND METHODS

Ethics statement

The study was approved by the Institutional Ethical Review Committee of the Sindh Institute of Urology and Transplantation, Karachi, Pakistan (Approval No. SIUT-ERC-2022/A-393). It was conducted in accordance with the ethical principles outlined in the modified Declaration of Helsinki.

Study population

A total of 329 adult patients aged ≥ 18 years, with biopsy-proven MPGN, were retrieved between January 2010 and December 2021. After excluding patients with secondary MPGN such as lupus nephritis (n = 27), post-infectious glomerulonephritis (n = 15), hepatitis B virus-associated glomerulonephritis (n = 13), and hepatitis C virus-associated glomerulonephritis (n = 17), as well as those with incomplete records or missing files (n = 100), those lost to follow-up before 6 months (n = 29), and those with pre-existing chronic kidney disease, 128 patients were found to be eligible. These cases were reclassified based on IF findings into IC-MPGN, defined as deposition of either IgG ≥ 1+ along with C3 and/or C1q or non-dominant IgA with C3 and/or C1q, and C-MPGN, which was diagnosed when C3 stained exclusively or predominantly (two orders of magnitude higher than any other IF reactant) on a scale of 0 to 3+ (Figure 1), according to the consensus guidelines of Kidney Disease: Improving Global Outcome[2].

Figure 1 Immunopathology of complement-mediated membranoproliferative glomerulonephritis A: Mesangial and endocapillary hypercellularity with lobular accentuation (hematoxylin and eosin, × 400); B: Mesangial proliferation with almost diffuse inter-positioning into peripheral capillary loops (periodic acid-Schiff stain, × 400); C: Mesangiocapillary proliferation with double contour formation exhibiting tram track appearance (silver methenamine stain, × 400); D: Diffuse, bright, ring-like positivity of C3 with an intensity of 3+ on a scale of 0-3, predominantly in the mesangium on immunofluorescence. The rest of the immunoreactions were two orders of magnitude lower in intensity (≤ 1+; immunofluorescence, × 400).

Data collection

Data were retrieved from the original renal biopsy request forms in the histopathology archives and corresponding case files. Information on demographics, clinical presentation, laboratory findings, treatment regimens, and outcomes was collected and analyzed. Clinical details included age, gender, systolic and diastolic blood pressure, initial clinical manifestations, treatment regimens administered, and final outcomes. Laboratory findings comprised serum creatinine (mg/dL), serum albumin (g/dL), and urine analysis reports (including hematuria and proteinuria) at presentation and during serial follow-ups. Serum C3 and C4 levels (recorded as low or normal) were also documented. The estimated glomerular filtration rate (eGFR) was calculated with the chronic kidney disease epidemiology collaboration creatinine equation[13]. Clinicopathological findings and factors influencing disease outcomes in the two groups were compared.

Evaluation of renal outcomes

Remission in idiopathic MPGN is categorized as complete remission, and partial remission, in accordance with Kidney Disease: Improving Global Outcomes guidelines[14]. The complete remission was defined as normal urinalysis (dipstick negative or trace for both proteins and blood), serum albumin > 3.5 g/dL with eGFR > 90 mL/minute/1.73 m². Partial remission was defined as abnormal urinalysis (microscopic hematuria or ≥ 1+ proteinuria), serum albumin < 3.5 g/dL, with eGFR < 90 mL/minute/1.73 m².

Renal survival was defined as the time from renal biopsy to the first of any of the following events: Starting dialysis, receiving a kidney transplant or an eGFR falling to < 15 mL/minute/1.73 m² at any point during follow-up and then not returning to > 15 mL/minute/1.73 m² at subsequent time points.

Evaluation of Histopathological findings

Histopathological evaluation included the total number of glomeruli and the percentage of sclerosed glomeruli, stratified as < 25%, 25%-50%, and > 50%. The presence of vasculopathy and crescents was noted, with crescents further classified as cellular or fibrocellular. Interstitial fibrosis and tubular atrophy (IFTA) were assessed and semiquantitatively graded as none (0), mild (< 25%), moderate (26%-50%), or severe (> 50%). Positive IF staining and the intensity for IgG, IgA, IgM, C3, and C1q were also recorded and subclassified as trace, +1, +2, or +3 (Figure 1).

Statistical analysis

Statistical analyses were performed using the Statistical Package for the Social Sciences, Version 22 (IBM Corporation, NY, United States). Categorical variables were expressed as n (%), while continuous variables were presented as mean ± SD or median (interquartile range). Differences between continuous variables were assessed using Student’s t-test (for normally distributed data) or the Mann-Whitney U test (for non-normally distributed data). Categorical variables were analyzed using the χ² test or Fisher’s exact test, as appropriate. Time-to-event analysis was performed using Kaplan-Meier survival curves, and the log-rank test was used to compare the results. Multivariate Cox regression analysis was done to identify independent risk factors for renal outcomes. A P < 0.05 was considered statistically significant.

RESULTS

Study design

This retrospective cohort study encompassed 128 adult patients (≥ 18 years) diagnosed with primary MPGN between January 2010 and December 2021. Upon reclassification, 100 patients (78.1%) were categorized as IC-MPGN, while the remaining 28 patients (21.9%) were categorized as C-MPGN.

Patient demographics, clinical, and laboratory parameters at presentation

Table 1 summarizes the demographic, clinical, and laboratory features of the two groups. At the time of biopsy, individuals with C-MPGN were significantly older than those with IC-MPGN (median age: 38 years vs 33.5 years; P = 0.029). Both groups demonstrated comparable systolic and diastolic blood pressures as well as similar initial clinical presentations. Serum creatinine tended to be higher and eGFR lower in C-MPGN, though the variation was not statistically significant. Nephrotic-range proteinuria was observed in both cohorts; however, IC-MPGN patients showed significantly reduced mean total serum protein (5.37 ± 1.03 vs 5.88 ± 0.99; P = 0.020) and albumin levels (2.40 ± 0.60 vs 2.68 ± 0.70; P = 0.036). Microscopic hematuria (≥ 1+) was also more prevalent in IC-MPGN (92%) compared with C-MPGN (75%; P = 0.018). Conversely, low serum C3 was detected more frequently in C-MPGN (82.1% vs 65%), though this difference did not reach statistical significance (P = 0.084).

Table 1 Comparison of baseline demographic, clinical, and laboratory features by immunofluorescence classification, median (interquartile rage)/n (%)/mean ± SD.

	IC-MPGN (n = 100)	C-MPGN (n = 28)	P value
Age at biopsy (years)	33.5 (22-45)	38 (28.3-53)	0.029
Gender			0.978
Male	64 (64.0)	18 (64.3)
Female	36 (36.0)	10 (35.7)
Blood pressure (mmHg)
Systolic	150 (135-160)	145 (139.2-159)	0.394
Diastolic	93.5 (80.5-100)	89 (80-100)	0.293
First clinical manifestation			0.298
Nephrotic syndrome	34 (34)	10 (35.7)
Acute GN	53 (53)	16 (57.1)
RPGN	13 (13)	2 (7.1)
Microscopic hematuria			0.018
< 2 (trace)	8 (8)	7 (25)
2-4 (+1)	17 (17)	3 (10.7)
5-10 (+2)	10 (10)	6 (21.4)
≥ 11 (+3)	65 (65)	12 (42.9)
Proteinuria (dipstick)			0.760
Trace	1 (1)	0 (0)
+1	3 (3)	0 (0)
+2	10 (10)	2 (7.1)
+3	65 (65)	18 (64.3)
+4	21 (21)	8 (28.6)
Serum creatinine (mg/dL)	2.25 (1.6-3.7)	2.55 (1.6-5.3)	0.537
eGFR (ml/minute/1.73 m2)	62.6 (40-108)	57 (20-99.8)	0.208
Total serum protein (g/dL)	5.37 ± 1.03	5.88 ± 0.99	0.020
Serum albumin (g/dL)	2.40 ± 0.60	2.68 ± 0.70	0.036
Serum C3 levels			0.084
Low (< 0.8 g/L)	65 (65)	23 (82.1)
Normal (> 0.8 g/L)	35 (35)	5 (17.9)
Serum C4 levels			0.965
Low (< 0.16 g/L)	29 (29)	8 (28.6)
Normal (> 0.16 g/L)	71 (70)	20 (71.4)
24 hours urinary protein (g/day)	3.1 (2-4.4)	3.01 (2-4.9)	0.753

Continuous variables were compared using the Mann-Whitney U test (or Student’s t-test for normally distributed data). Categorical variables were analyzed using χ² or Fisher’s exact test, as appropriate. GN: Glomerulonephritis; RPGN: Rapidly progressive glomerulonephritis; eGFR: Estimated glomerular filtration rate; IC-MPGN: Immune complex-mediated membranoproliferative glomerulonephritis; C-MPGN: Complement-mediated membranoproliferative glomerulonephritis.

Histopathological findings

Table 2 outlines the histopathological features observed on renal biopsies in both cohorts. The median glomerular count per biopsy was identical in IC-MPGN and C-MPGN (16 each; P = 0.924). The prevalence of global glomerulosclerosis also showed no significant variation (P = 0.725), with the majority of samples in both groups containing ≤ 25% sclerosed glomeruli. Crescents were more common in C-MPGN (60%) than IC-MPGN (48%), mainly of the cellular type, though the difference lacked statistical significance (P = 0.385). IFTA was generally mild across the cohorts, with a greater share of C-MPGN cases demonstrating absent or minimal changes compared with IC-MPGN (89.2% vs 78%; P = 0.270), but without statistical relevance. Notably, vascular pathology was significantly more frequent in C-MPGN (50.0%) compared with IC-MPGN (28.0%; P = 0.028).

Table 2 Comparison of histopathological characteristics between the two groups of membranoproliferative glomerulonephritis classified based on immunofluorescence results, median (interquartile rage)/n (%).

	IC-MPGN (n = 100)	C-MPGN (n = 28)	P value
Total glomeruli	16 (11-23)	16 (12-20)	0.924
Global sclerosis			0.725
None	54 (54)	14 (50)
25%	30 (30)	11 (39.3)
26%-50%	13 (13)	2 (7.1)
> 50%	3 (3)	1 (3.6)
Vasculopathy	28 (28)	14 (50)	0.028
Crescents			0.385
None	52 (52)	11 (39.3)
Cellular	39 (39)	15 (53.6)
Fibrocellular	9 (9)	2 (7.1)
IF/TA			0.270
None	15 (15)	2 (7.1)
Mild	63 (63)	23 (82.1)
Moderate	19 (19)	3(10.7)
Severe	3 (3)	0 (0.0)
Immunofluorescence results
IgG			0.000
Negative	11 (11)	17 (60.7)
Trace	2 (2)	3 (10.7)
+1	77 (77)	8 (28.6)
+2	9 (9)	0 (0)
+3	1 (1)	0 (0)
IgA			0.275
Negative	84 (84)	28 (100)
Trace	1 (1)	0 (0)
+1	12 (12)	0 (0)
+2	1 (1)	0 (0)
+3	2 (2)	0 (0)
IgM			0.001
Negative	56 (56)	26 (92.9)
Trace	0 (0)	0 (0)
+1	36 (36)	2 (7.1)
+2	8 (8)	0 (0)
+3	0 (0)	0 (0)
C3			0.000
Negative	10 (10)	0 (0)
Trace	4 (4)	0 (0)
+1	53 (53)	0 (0)
+2	27 (27)	9 (32.1)
+3	6 (6)	19 (67.9)
C1q			0.039
Negative	64 (64)	15 (53.6)
Trace	0 (0)	1 (3.6)
+1	24 (24)	12 (42.9)
+2	9 (9)	0 (0)
+3	3 (3)	0 (0)

Continuous variables were compared using the Mann-Whitney U test (or Student’s t-test for normally distributed data). Categorical variables were analyzed using χ² or Fisher’s exact test, as appropriate. IF: Interstitial fibrosis; TA: Tubular atrophy; IC-MPGN: Immune complex-mediated membranoproliferative glomerulonephritis; C-MPGN: Complement-mediated membranoproliferative glomerulonephritis.

IF findings

Table 2 also highlights clear distinctions in IF patterns between the two groups. In C-MPGN, IgG was absent or trace in over 70% of cases, whereas 77% of IC-MPGN biopsies showed 1+ staining, a difference that was highly significant (P < 0.001). IgM was similarly scarce in C-MPGN (93% negative) compared with IC-MPGN (56% negative; P = 0.001). By contrast, C3 deposition was markedly stronger in C-MPGN (P < 0.001), with all cases demonstrating 2+ to 3+ positivity, while only one-third of IC-MPGN biopsies reached this level. C1q was consistently ≤ 1+ in C-MPGN (100% vs 88% in IC-MPGN; P = 0.039), and 12% of IC-MPGN samples showed ≥ 2+ deposition. IgA staining did not differ significantly (P = 0.275), with most biopsies in both groups remaining negative.

Renal outcomes

Table 3 illustrates a comparison of renal function and outcomes over 36 months in IC-MPGN and C-MPGN. At baseline, the median eGFR was 62.6 mL/minute/1.73 m² in IC-MPGN compared with 57 mL/minute/1.73 m² in C-MPGN, with values rising progressively in both groups during follow-up. By 36 months, eGFR remained slightly higher in IC-MPGN (72.2 mL/minute/1.73 m²) than in C-MPGN (68 mL/minute/1.73 m²), though the difference was not statistically significant (P = 0.62). Serum albumin also improved gradually in both cohorts without significant differences. The clinical response rates were higher in IC-MPGN, with 62% achieving complete or partial remission compared with 60.7% in C-MPGN (log-rank test: P = 0.40). Kaplan-Meier survival analysis was performed by considering the period from treatment initiation to the end of follow-up or death as the time frame (Figure 2). Progression to ESKD was similar between the two groups, as was dialysis dependence (26% vs 25%; P = 0.55). Mortality was slightly increased in IC-MPGN [4 cases (4%) vs 0], though this difference did not reach statistical significance (P = 0.24). All deaths were related to infectious complications, and the absence of mortality in the CMPGN group likely reflects smaller sample size and followup limitations rather than true differences in the rates.

Figure 2 Kaplan-Meier survival analysis curves. A comparison of survival rates between immune complex-mediated membranoproliferative glomerulonephritis and complement-mediated membranoproliferative glomerulonephritis patients analyzed using the log-rank test. IC-MPGN: Immune complex-mediated membranoproliferative glomerulonephritis; C-MPGN: Complement-mediated membranoproliferative glomerulonephritis.

Table 3 Renal function comparison between immune complex-mediated membranoproliferative glomerulonephritis and complement-mediated membranoproliferative glomerulonephritis at baseline and 36 months, including outcomes, median (interquartile rage)/n (%)/mean ± SD.

	IC-MPGN (n = 100)	C-MPGN (n = 28)	P value
At presentation
Serum creatinine (mg/dL)	2.25 (1.6-3.7)	2.55 (1.6-5.3)	0.537
eGFR (mL/minute/1.73 m2)	62.6 (40-108)	57 (20-99.8)	0.208
Serum albumin (g/dL)	2.40 ± 0.60	2.68 ± 0.70	0.036
Patients followed for 36 months	88 (88)	24 (85.7)
Serum creatinine (mg/dL)	1.5 (0.80-7.0)	1.8 (1.0-7.7)	0.507
eGFR (mL/minute/1.73 m2)	72.2 (14-118)	68 (22-95.2)	0.62
Serum albumin (g/dL)	3.17 ± 0.76	3.32 ± 0.79	0.385
Final outcomes			0.555
CR	27 (27)	5 (17.85)
PR	35 (35)	12 (42.85)
ESKD	26 (26)	7 (25)
Lost to FU	8 (8)	4 (14.2)	0.32
Overall mortality	4 (4)	0 (0)	0.24
Renal survival	62 (62)	17(60.7)	0.62

Continuous variables were compared using the Mann-Whitney U test (or Student’s t-test for normally distributed data). Categorical variables were analyzed using χ² or Fisher’s exact test, as appropriate. eGFR: Estimated glomerular filtration rate; CR: Complete remission; PR: Partial remission; ESKD: End-stage kidney disease; FU: Follow-up; IC-MPGN: Immune complex-mediated membranoproliferative glomerulonephritis; C-MPGN: Complement-mediated membranoproliferative glomerulonephritis.

Although remission rates differed numerically (62% vs 60.7%), posthoc power analysis of the primary outcome comparison showed that, with the current sample sizes (ICMPGN n = 100, CMPGN n = 28), the achieved power was only 12% at α = 0.05. The corresponding effect size (Cohen’s h) was 0.03, indicating a very small difference. Detecting such a difference with 80% power would require approximately 1400 patients per group. This limitation underscores that the present study was underpowered to detect subtle differences, highlighting the need for larger, multicenter cohorts to validate these findings and better assess subgroup outcomes.

Treatment regimens and outcomes

A higher proportion of C-MPGN patients received immunosuppressive therapy compared with IC-MPGN (67.8% vs 51%). Steroid monotherapy was administered to 35 individuals with IC-MPGN (35%) and 8 with C-MPGN (28.6%), whereas combination therapy was more common in C-MPGN (39.3% vs 16%). At 36 months, overall remission rates were similar between groups, with 56.9% of IC-MPGN and 57.9% of C-MPGN patients achieving complete or partial remission (P = 0.68). However, complete remission was observed only in IC-MPGN (22.9%), with none of the C-MPGN cases achieving CR under steroid therapy (P = 0.134). Progression to ESKD and dialysis dependence was slightly higher in C-MPGN [8 (42.1%)] compared with IC-MPGN [18 (35.3%); P = 0.68]. Mortality occurred exclusively in IC-MPGN, with 4 deaths (4%), while no deaths were reported in C-MPGN (Table 4).

Table 4 Therapeutic strategies, treatment response, and renal outcomes in immune complex-mediated membranoproliferative glomerulonephritis vs complement-mediated membranoproliferative glomerulonephritis by immunosuppressive regimen, n (%).

	IC-MPGN (n = 100)	C-MPGN (n = 28)	P value
Immunosuppressives	51 (51)	19 (67.8)
Steroids alone	35 (35)	8 (28.57)
CR	8 (22.9)	0 (0)	0.134
PR	13 (37.1)	4 (50)	0.692
ESKD	12 (34.2)	4 (50)	0.68
Mortality	2 (5.7)	0 (0)	0.99
Cyclophosphamide + steroids	16 (16)	11 (39.28)
CR	2 (12.5)	3 (27.3)	0.37
PR	6 (37.5)	4 (36.4)	0.99
ESKD	6 (37.5)	4 (36.4)	0.89
Mortality	2 (12.5)	0 (0)	0.49

Continuous variables were compared using the Mann-Whitney U test (or Student’s t-test for normally distributed data). Categorical variables were analyzed using χ² or Fisher’s exact test, as appropriate. CR: Complete remission; PR: Partial remission; ESKD: End-stage kidney disease; IC-MPGN: Immune complex-mediated membranoproliferative glomerulonephritis; C-MPGN: Complement-mediated membranoproliferative glomerulonephritis.

Factors associated with disease remission and ESKD

By performing multivariate Cox regression analysis, we examined the factors affecting remission rates and ESKD between the two groups. In the IC-MPGN group, no variables were independently associated with kidney survival in terms of remission; however, moderate-to-severe tubular atrophy emerged as an independent predictor of poor kidney outcomes. Similarly, in the C-MPGN group, no variables were independently associated with kidney survival, while vasculopathy and the presence of crescents were identified as independent predictors of ESKD (Table 5).

Table 5 Cox regression analysis for factors associated with disease remission and end-stage kidney disease among the two groups of membranoproliferative glomerulonephritis.

Variables	Remission		ESKD		Remission		ESKD
	HR (95%CI)	P value	HR (95%CI)	P value	HR (95%CI)	P value	HR (95%CI)	P value
Blood pressure on admission	1.01 (0.34-1.46)	0.72	1.12 (0.71-1.8)	0.79	0.39 (0.11-1.16)	0.07	0.45 (0.22-1.8)	0.63
Proteinuria on admission (g/day)	1.25 (0.75-2.48)	0.29	0.84 (0.32-1.96)	0.89	0.38 (0.14-1.10)	0.06	0.53 (0.29-1.1)	0.54
Vasculopathy	0.97 (0.67-1.8)	0.07	1.58 (0.6-1.89)	0.62	0.43 (0.26-1.17)	0.24	1.2 (0.50-3.10)	0.04
Presence of crescents	0.27 (0.16-0.8)	0.07	0.49 (0.3-0.81)	0.32	1.02 (0.22-1.8)	0.08	1.3 (0.23-3.2)	0.02
IF/TA	1.1 (0.58-1.6)	0.24	1.45 (1.1-3.2)	0.02	1.9 (0.62-5.1)	0.19	1.5 (0.82-6.01)	0.07
Steroids alone	0.67 (0.54-2.1)	0.42	0.28 (0.16-1.27)	0.08	0.29 (0.16-1.15)	0.07	1.39 (0.16-4.01)	0.71
Cyclophosphamide	1.21 (0.48-2.75)	0.49	0.78 (0.2-1.8)	0.71	0.9 (0.30-2.0)	0.87	0.22 (0.23-1.96)	0.26

IF: Interstitial fibrosis; TA: Tubular atrophy; HR: Hazard ratio; CI: Confidence interval; ESKD: End-stage kidney disease.

DISCUSSION

This study provides a detailed comparison of IC-MPGN and C-MPGN, classified according to up-to-date IF criteria. In this study, we compared the clinical, laboratory, and histopathological features and renal outcomes of patients with IC-MPGN and C-MPGN, providing a longitudinal analysis for up to 36 months. Our findings highlight important distinctions in pathophysiology, clinical presentation, and response to therapy, while demonstrating broadly similar renal outcomes over time, the latter partly because of unbalanced sample sizes and differential loss-to-follow-up.

In this study, patients were mostly young, with only modest age difference between the two groups, which is consistent with findings from previously published studies[15]. Patients with C-MPGN were slightly older at the time of biopsy (median age: 38 years) compared with IC-MPGN (33.5 years). This contrasts with another study that documented a marked difference in mean ages between the two subtypes (62 years vs 19 years)[16]. Across populations, however, no consistent age disparity has been noted between Asian and Western cohorts. Gender distribution was comparable in both groups, consistent with previous findings that show no clear gender predominance[17].

There remains ongoing debate concerning the extent to which the baseline clinical and histopathological features can predict long-term outcomes in MPGN[18]. Previous studies conducted before the current reclassification identified older age, hypertension, nephrotic-range proteinuria, reduced eGFR, extensive glomerulosclerosis, crescent formation, and marked interstitial fibrosis as harbingers of poor prognosis[19-23]. In our cohort, most patients presented with acute glomerulonephritis followed by nephrotic syndrome, a pattern that contrasts with earlier reports describing a predominance of nephrotic syndrome in IC-MPGN[15,16]. Proteinuria and renal function at baseline were largely comparable between the two groups, aligning with observations by Sethi and Fervenza[1] and Servais et al[24], who emphasized that renal function at presentation does not reliably differentiate IC-MPGN from C-MPGN, emphasizing the diagnostic value of renal biopsy and IF[1,2]. Notably, patients with IC-MPGN in our study exhibited significantly lower mean total serum protein and albumin levels, along with more pronounced hematuria compared to those with C-MPGN, consistent with the findings reported by Elahi et al[25].

Furthermore, in our cohort, the overall histopathological features of IC-MPGN and C-MPGN were largely comparable to those of Nakano et al[15] and Nakagawa et al[16]. The number of glomeruli obtained per biopsy and the extent of global glomerulosclerosis showed no significant differences between the two groups, with most samples demonstrating only limited sclerosis. Crescent formation was observed more frequently in C-MPGN, predominantly of the cellular type, though this difference did not reach statistical significance. IFTA was generally mild in both groups, with C-MPGN cases more often showing absent or minimal changes, again without significant differences. A notable finding was the higher prevalence of vascular pathology in C-MPGN compared with IC-MPGN. This observation contrasts with the studies by Kovala et al[26,27], which reported no differences in global sclerosis, crescents, or IFTA between the two subtypes but did not emphasize vasculopathy. Our results, therefore, suggest that complement-mediated injury may involve the microvasculature more prominently than immune complex deposition, supporting previous reports that highlight vascular involvement as a distinguishing feature of C-MPGN.

The IF patterns in our cohort were in congruence with the established classification criteria and provided a clear distinction between the two entities. IC-MPGN was marked by prominent IgG and IgM deposition, whereas C-MPGN demonstrated intense C3 staining with minimal immunoglobulin, reflecting the central role of alternative pathway dysregulation in the latter condition[4,7]. This reinforces the pivotal role of IF microscopy in accurately subclassifying MPGN and informing therapeutic choices. Moreover, the pronounced C3 deposition in C-MPGN was accompanied by markedly reduced serum C3 Levels, indicative of enhanced complement consumption and its contribution to disease progression.

The overall renal prognosis in adult MPGN is generally poor, as highlighted in several studies[28,29], including work from Korea, where MPGN was associated with the worst outcomes among primary glomerulonephritides[30]. However, many of these studies were limited by small cohorts, inclusion of mixed adult and pediatric populations, or failure to exclude secondary causes[15,19-20]. Findings across the literature remain inconsistent, with some studies reporting a less favorable prognosis in IC-MPGN compared with C-MPGN[19,20], while others suggest the opposite[16], and several showing no clear differences[15,21,22,31]. In contrast, pediatric patients with IC-MPGN and C-MPGN often show a more favorable course, with preserved renal function and rare progression to advanced chronic kidney disease. This likely reflects the longer time required to reach ESKD or mortality endpoints, as well as the shorter duration of published follow-up[21,23,31]. More recent data illustrate the variability in outcomes: Bomback et al[32] reported substantial progression to ESKD or death in C-MPGN within five years, while Nakano et al[15] observed progression in both IC-MPGN and C-MPGN over longer follow-up. In our study, three-year renal survival was comparable between the two cohorts, approximating 60%-62%, a finding that contrasts with results from South Africa, where survival in IC-MPGN was notably lower[33].

Management of MPGN remains complex owing to its heterogeneity and the lack of standardized treatment guidelines. Steroids continue to serve as the mainstay of therapy, yet patient responses are highly variable, ranging from favorable outcomes in some reports[17] to poor[34] or inconclusive results in others[15]. Evidence supporting cyclophosphamide use largely predates the current classification[35], and while certain studies suggested benefit, our cohort did not demonstrate improvement in renal outcomes with steroid-cyclophosphamide regimens. Similarly, although mycophenolate mofetil has been associated with remission in some series, other investigations have reported equivocal results[36,37], and no patients in our study received mycophenolate mofetil. Overall, treatment responses did not differ significantly between IC-MPGN and C-MPGN, underscoring the limitations of conventional immunosuppression. The variability and often suboptimal efficacy of these regimens highlight the unmet need for more targeted approaches. Complement-directed therapies, particularly anti-C5 monoclonal antibodies such as eculizumab, have shown promise in select cases of C-MPGN, though outcomes remain inconsistent[38]. As research advances, the therapeutic landscape is expected to evolve substantially, offering more precise and durable disease control for these challenging glomerulopathies. For this, early and non-invasive diagnostic biomarkers for this disease and underlying co-morbidities need to be investigated and adopted to achieve the best renal and patient outcomes[39,40].

In summary, the reclassification of MPGN has bridged a considerable knowledge gap in determining the most effective therapeutic strategies under the revised framework. In this study, outcomes between IC-MPGN and C-MPGN revealed no statistically significant differences in treatment response or renal prognosis following either steroid monotherapy or combined steroid-cyclophosphamide therapy. Although complete and partial remission rates were numerically higher in IC-MPGN, the persistent risk of progression to ESKD remained, and these variations did not achieve statistical significance, likely reflecting the limited sample size. Mortality was infrequent across the whole study cohort. Only four patients died in the IC-MPGN group and none in the C-MPGN group. The deaths in the ICMPGN group were related to infectious complications rather than treatmentrelated or cardiovascular causes. No specific explanation can be provided for the absence of mortality in the CMPGN group, which may reflect the small sample size and limited followup. The absence of definitive statistical differences in therapeutic responses and outcomes limits firm conclusions, highlighting the need for larger, multicenter investigations to establish optimal immunosuppressive approaches for these uncommon glomerulopathies. Future studies should also prioritize the evaluation of complement-directed therapies, which hold promise for delivering more targeted and sustained disease control, particularly in C-MPGN.

This study has several limitations. Firstly, the retrospective, single-center design introduces potential selection bias, missing data, and limits generalizability to broader populations. Secondly, the relatively small sample size, particularly for the C-MPGN subgroup, together with the restricted follow-up period of 36 months, reduced statistical power and precluded robust subgroup comparisons or the assessment of long-term outcomes such as ESKD. Thirdly, electron microscopy was not performed in all cases, preventing further subclassification of C3G into C3GN and DDD, which may carry different prognostic implications. Fourth, complement functional assays, genetic testing, extended autoantibody evaluation, and systematic screening for dysproteinemia or monoclonal gammopathy were not undertaken, restricting insights into underlying pathophysiology. Fifth, treatment regimens were not standardized across patients, reflecting the absence of universal protocols and potentially influencing therapeutic outcomes. Sixth, differential loss to followup was observed, with 8% of IC-MPGN patients and 14.2% of C-MPGN patients lost to followup, representing an approximately 76% higher attrition rate in the latter group. This imbalance could introduce survival bias and should be considered when interpreting outcomes. Finally, of the 329 patients initially identified, 201 (61%) were excluded, 101 due to secondary causes and 100 because of missing data. Given the limited demographics available, we cannot determine whether excluded patients differed systematically from those included. This high exclusion rate may have introduced bias and represents an important limitation of the study.

CONCLUSION

The majority of MPGN cases were reclassified as IC-MPGN, which showed numerically higher remission rates. Overall, the clinicopathological features at baseline were similar between the two groups, though C-MPGN, a less frequent subtype, was associated with vascular pathology and stronger complement deposition, with a poorer medium-term prognosis. Our findings reinforce the need for accurate subclassification using IF and highlight the potential utility of targeted complement-directed therapies, particularly for C-MPGN. Multicenter prospective studies with standardized treatment protocols and comprehensive complement analysis are warranted to optimize management and long-term prognosis.