Crescentic glomerulonephritis: what’s different in South Asia? A single center observational cohort study

Background: The spectrum and outcomes of crescentic glomerulonephritis (Cr.GN) in South Asia is vastly different from that reported worldwide and there is a paucity of information. The aim of the study was to study the demography, clinical presentation, histology and predictors of longitudinal outcomes of Cr.GN in this population. Methods: An observational cohort study of renal biopsies was performed in the largest tertiary center in South India over a period of 10 years (January 2006 to December 2015) with ≥50% crescents on renal histology indicating Cr.GN. Results: A total of 8645 kidney biopsies were done; 200 (2.31%) were Cr.GN. Patients were categorized into three etiological groups: anti-glomerular basement membrane (type I), immune complex (type II), and pauci-immune (type III). Type II was the most common (96, 46.5%), followed by type III (73, 38%) and type I (31, 15.5%). Female preponderance was seen across all types. About half of all patients presented with recent onset hypertension. Type II had the highest median proteinuria (4.2 (2.1-6) g/day, p=0.06) and the median estimated glomerular filtration rate was lowest in type I (5 (4-8) ml/min/1.73m 2, p<0.001). Among type III, anti-neutrophil cytoplasmic antibodies (ANCA)-associated vasculitis was seen only in ~50% of patients. Nearly one third of patients with type I were also positive for ANCA making them ‘double positive’. Acute glomerular insults like tuft necrosis and chronic changes as evidenced by moderate to severe interstitial fibrosis, was a predominant feature of type I. Conclusions: ANCA-negative pauci-immune vasculitis, as well as double positive Cr.GN, are reported for the first time in South-Asia. Renal survival was significantly worse in type I/III compared to type II. Types I/III, moderate to severe interstitial fibrosis and tubular atrophy, presence of oliguria/anuria and increasing percentage of crescents in renal biopsy were significant predictors of end stage kidney disease in our cohort.


Introduction
Crescentic glomerulonephritis (Cr.GN) is defined histologically by the presence of extensive glomerular crescents (usually greater than 50%). Clinically, it is also known as rapidly progressive glomerulonephritis (RPGN) as it is accompanied by rapid decline in renal functions. It can occur in any glomerular disease 1 and is usually reported in about 4% to 10% of native kidney biopsies 2 . The natural course of the disease is akin to "medical emergency", as end stage kidney disease (ESKD) is reached in most patients within a few weeks to months 3 . Understanding the clinical presentation, natural history and outcomes of Cr.GN is of major concern for nephrologists worldwide. Various studies have been conducted worldwide and epidemiologic data are available from large national kidney biopsy registries, including those from the United States 4 , China 2 , Japan 5 , Spain 6 and Saudi Arabia 7 . Notably, pathogenesis of glomerular diseases involves a complex and as yet incompletely understood interplay between epigenetic, immunoregulatory, hormonal, and environmental factors on a background of genetic predisposition 8 . This translates into a broad spectrum of disease presentation, a variable tempo of progression and heterogenous outcomes, which are evident from these previous studies. To this end, exploring the features of Cr.GN in a South Asian population, a genetically and demographically diverse group of individuals, may help to provide useful insights into the causality, and predictors of severe outcomes. Our aim was to study the demography, clinical presentation, histology and predictors of longitudinal outcomes of Cr.GN in this population.

Study design, setting and participants
This was an observational retrospective cohort study performed at the outpatient and inpatient services of the Department of Nephrology, Christian Medical College Vellore, India.
We included all patients (≥18 years) who underwent native renal biopsy at our centre between January 2006 to December 2015 and had ≥50% crescents on renal histology. There were no other inclusion or exclusion criteria.

Data collection
Data on patients' demographic profile, clinical features, biochemical parameters, histopathology, treatments, morbidity and mortality were retrieved from the electronic patient records (Clinical WorkStation) maintained in the hospital. Follow-up clinical and outcome data with regards to their serum creatinine, dialysis requirement, and complications were collected for each follow-up visit until August 2016. During the index and followup visits, patients were classified into chronic kidney disease (CKD) stages as per estimated glomerular filtration rate (eGFR) calculated by CKD-EPI equation 9 . Quantitative determination of complement factors (C3 and C4) was done by means of endpoint nephelometry on the BN ProSpec System by Siemens Health Care Diagnostics Products, Marburg, Germany. Antisera used were liquid animal sera produced by immunization of rabbits with highly purified human complement factors (C3c or C4). The following reference intervals applied for serum samples from healthy adults: C3/C3c, 0.9-1.8 g/L; C4/C4c, 0.1-0.4 g/L.

Statistical analysis
Data are presented as mean ± standard deviation or medians (interquartile range) or frequency and percent (%) according to the types and distribution of variables. Differences among groups of normally distributed variables were analyzed by t test or one-way analysis of variance (ANOVA  Figure 1. Females constituted 60% of the patients with a female: male ratio of 1.5:1. Female preponderance was seen across all three types of Cr.GN. The mean age of presentation for all types was 40.6±14.6 years, with the highest mean age of presentation seen in patients with type III Cr.GN. Demographic and baseline clinical and laboratory parameters of the study population are summarized in Table 1.

Clinical and laboratory features
Non-visible hematuria was near universal (95%). More than half of the patients (56%) were oliguric at presentation. Anuria at presentation was seen in 10% patients. Oliguria and anuria were more common in type I Cr.GN patients (oliguria in 74%, p=0.08; anuria in 16%, p=0.04) who also had significantly more uremic symptoms (64%, p=0.006). About half of all the three types presented with recent onset hypertension. Among extra-renal symptoms, skin lesions and arthritis were rare in type I, but hemoptysis was seen only in type I and III Cr.GN.

Treatment differences
Standard treatment protocols as per KDIGO 2012 10 guidelines were followed for all patients. Nearly half of the patients required dialysis at presentation, with significantly more in type I and type III Cr.GN (81% & 62% respectively, p<0.001).
About one fifth of patients received therapeutic plasma exchange (PLEX), and this was significantly more in type I and type III Cr.GN (42% and 26% respectively, p<0.001). The frequency and indications of PLEX are summarized in Table 3. Immunosuppression with/without PLEX was given in 90% of patients with significantly more in type II and III Cr.GN (92% and 91% respectively, p<0.001).

Patient and renal outcomes
The mean follow-up period was 9.4±15 months. Table 4 summarizes the outcomes observed in three types of Cr.GN. In the entire cohort, nearly half (45%) developed ESKD, requiring renal replacement therapy. This was significantly more in type I (77%, p<0.001). Overall mortality rate was 4% and did not vary significantly between the groups. Sepsis was found to be the most common cause of death.

Predictors of severe outcomes
We analyzed various risk factors which predicted the development of ESKD at follow-up in this cohort (Table 6). In an adjusted regression analysis, type I/type III Cr.GN, moderate to severe IFTA, presence of oliguria/anuria and increasing percentage of crescents in renal biopsy were significant predictors of ESKD at follow-up.

Discussion
Cr.GN is one of the leading causes of rapidly progressive renal failure. There are a few studies of Cr.GN from South Asia but there is dearth of data on the different types of Cr.GN and their outcomes.
In this study Cr.GN accounted for 2.3% of all biopsies conducted over a period of 10 years, which is comparable to previously reported rates of Cr.GN from India 11,12 . Type II Cr.GN was found to be the most common type, which is unique to the Asian continent. Various studies conducted in different parts of the world have shown type III Cr.GN to be the most common 5,13 . However studies from China, Saudi Arabia and a smaller Indian study have reported type II Cr.GN to be the most common 2,11,14 . Reasons cited for increased incidence of type II Cr.GN in these parts have been an increased incidence of infections and a higher prevalence of IgA nephropathy. Lupus nephritis (45%) is the most common cause of type II Cr.GN, followed by infection related GN (24%) and IgA nephropathy (23%) in our cohort. This is similar to one of the largest reviews of Cr.GN from China, in which lupus nephritis was listed as most common cause of type II Cr.GN (34%) followed by IgA nephropathy (17%) 2 .
Gender differences in Cr.GN have mostly been observed in type II Cr.GN with female preponderance. The gender distribution has been variable in type I and type III Cr.GN in different studies 2,11,13-15 . As is already known and established in other studies, patients with type I Cr.GN had the most severe renal failure at presentation. However, in this study, more than half of the patients with type II and III Cr.GN also presented with severe renal failure, highlighting the dramatic presentation in this patient population. These rates are much higher than previously reported 2,15 .
Serum complement C3 levels have been shown to be low in type I and type III Cr.GN in addition to type II Cr.GN. This was confirmed in our study. However, we found no cases of low C4 levels in type I and III Cr.GN, highlighting that the alternate complement pathway has a role in the pathogenesis of these GNs. Our observed rates of ANCA seropositivity in type III Cr.GN are much lower than those reported in other studies 2,15 . An earlier study from India also reported similar rates of ANCA seropositivity 16 . ANCA negative vasculitis could be attributed to other antibodies, such as anti-endothelial cell antibodies, or to cell mediated immune mechanisms, which lead to neutrophilic activation 17 . The high prevalence of ANCA negative vasculitis in this cohort highlights the need for research into its pathogenesis to elucidate factors specific to our population.
Subgroup analysis of type III Cr.GN revealed important differences between AAV and ANCA negative vasculitis groups, which, to the best of our knowledge, is reported for the first time from India. Similar to our study, younger age of onset and lower prevalence of systemic involvement in ANCA negative vasculitis has been reported from other parts of the world 18-21 . Chen et al. 19 also observed a higher level of proteinuria in this group, similar to our study. Although chronic lesions on kidney biopsy were more prevalent in ANCA negative vasculitis in these studies, we found a higher prevalence of fibrous/fibrocellular crescents and tuft necrosis in the AAV group.

Conclusion
We were able to analyse detailed demographic, clinical, serological and pathological features of our cohort. Strengths of our study include its large sample size, the inclusion and comparison of ANCA negative vasculitis as well as 'double positive Cr.GN' patients, of which there is no previously reported data from India. We highlight several important clinical practice points, in particular type II Cr.GN may present with a severe renal failure similar to type III Cr.GN. However, the response to treatment and outcomes were much more favourable and appropriate treatment should be initiated at the earliest. Prevalence of ANCA negative vasculitis was much higher in our population and hence kidney biopsy is mandatory in a case of suspected RPGN with negative serologies. Type I/type III Cr.GN, moderate to severe IFTA, presence of oliguria/anuria, and increasing percentage of crescents in renal biopsy were significant predictors of ESKD at follow-up in our cohort. Our study has also been able to identify areas of further research: search for pathways of alternate complement activation in type I and III Cr.GN and to explore the pathogenesis and factors responsible for increased prevalence of ANCA negative vasculitis in our population. The limitations of the study are that it is a single centre and it was retrospective in nature.  The frequency of this disease amongst biopsied patients with glomerulonephritis (2.3%) is a lower frequency than has been found in other large studies. However this was a relatively large group (200) enabling more confidence in the data collected.

Data availability
Relatively to other large studies there were several significant observations. The relative frequency of infection as an underlying disease was unusually high. There was also a relatively higher than expected frequency of Lupus Nephritis.
The other unexpected findings were the high percentage of ANCA negative patients amongst the Type III pauci immune group. These two groups, (Type III ANCA positive and negative), were similar in most clinical and biochemical features except for the observation that ANCA negative patients were significantly younger, had significantly less extra renal disease especially haemoptysis. Consequently they had significantly less plasma exchange treatment. Their biopsies showed less glomerulosclerosis and full glomerular tuft necrosis and they had a strong trends to outcomes with less loss of renal function and death.
The frequency of Type I, anti GBM disease group was, as expected, infrequent. Only 21 patients had "pure" anti-GBM disease but the number of double positive (10 patients with Type I anti GBM disease and concurrently Type III classification) significantly raised the percentage of patients with anti GBM disease. If we deleted these 10 double positive patients from the Type III group this "Pure Type III" group would now fall to 68 and the number of ANCA positive patients in this "Pure Type III group" would fall to 28. Therefore the percentage of "pure Type III "patients ANCA positive would fall to 41%. This is a remarkably low frequency of ANCA positivity in Type III Cr.GN patients. It would be interesting to see how many of these patients ("Pure" anti GBM disease, "Pure Type III and" double positive) had circulating anti-GBM antibodies.