Membranous glomerulonephritis: Role of Retinol-binding Protein in monitoring and prognostication

Retinol binding protein (RBP) which was initially thought to be a biomarker for proximal convoluted tubule dysfunction, could be important in chronic kidney disease. Membranous glomerulonephritis (MGN) was considered one of the most common causes of chronic kidney diseases (CKD) and subsequent progression to End-stage Renal Disease (ESRD). As such, monitoring MGN subjects especially those at risk of progression to ESRD is quite important for prompt intervention and treatment decision. This study aimed at using a non-invasive biomarker (urine RBP) for effective monitoring and prognostication of MGN subjects using the Enzyme-linked Immunosorbent Assay (ELISA) method. One hundred and twenty-five, including 81 primary and 44 secondary MGN subjects, were diagnosed with MGN from January 2012 to October 2019 at Hospital Serdang and Hospital Kuala Lumpur from which 69 subjects consisting of 45 primary and 24 secondary MGN subjects participated in the study. The test for biomarkers using the ELISA technique gave the following results: urine RBP was detected in 27 (39.1%) and 6 (8.7%) primary and secondary MGN subjects respectively. Correlation analysis shows a significant correlation between urinary RBP and renal function test parameters, in addition to logistic regression analysis which demonstrated that urinary RBP is a prognostic biomarker for primary MGN. Therefore, urinary RBP could be used in monitoring and prognostication of primary MGN subjects.


Introduction
Retinol binding protein is a low molecular weight protein, synthesized in the liver (1), binds to transthyretin (a complex that prevents glomerular filtration), and is found in plasma circulation (2). The principal function of RBP is vitamin A transport (3). It has been documented that about 4-5% of RBP circulates freely, crosses the glomerular barrier and is Preprints (www.preprints.org) | NOT PEER-REVIEWED | Posted: 4 August 2021 doi:10.20944/preprints202108.0109.v1 reabsorbed at the proximal convoluted tubule (PCT) (4). Individualized therapy is possible with the development of a non-invasive biomarker such as urinary RBP to monitor chronic kidney disease (CKD) patients at risk of progressing to end-stage renal disease (ESRD) (5,6). Besides, urinary RBP is linked to a high risk of creatinine doubling and renal replacement therapy initiation in CKD (7). Therefore, urinary RBP could serve as a non-invasive biomarker for PCT dysfunction (8) and CKDs (4,9,10) such as diabetes mellitus (DM), HIV-associated nephropathy (HIVAN) for effective intervention (11). Membranous glomerulonephritis (MGN) is one of the leading causes of CKD and progression to ESRD is linked to high morbidity and mortality (4,12). Therefore, early intervention and monitoring are necessary to prevent patients from progressing to ESRD. Previous researches demonstrated that higher urinary RBP is related to the possibility of ESRD and corticosteroid tolerance in acute glomerulonephritis (13,14). Low serum albumin was defined as < 35g/L, serum creatinine as 44-80mol/L and serum urea as 2.76-8.07mmol/L. The primary outcomes were defined as eGFR < 60 ml/min/1.73m 2 (CKD≥ 3) and UPCr Index > 0.03 g/mmol (no remission)

Materials and Method
while the reference value for urinary RBP was 0.86 pg/mL (optimized by ROC curve).

Analysis of data
The statistical analysis was performed through version 25.0 of the standard software package (SPSS). The mean, the standard deviation was used for normally distributed variables, while the median (interquartile) was used for variables that are not normally distributed. Categorical variables were represented as frequencies and percentages and compared by chi-square. Simple and multiple regressions were conducted to determine the primary outcome, and the difference was found to be statistically significant at p< 0.05.

Characteristics of the patients at presentation (n= 69)
Sixty-nine patients, including 47 primary MGN and 22 secondary MGN, were involved in this research.

Relationship between urinary RBP and renal function test parameters at the end of the follow-up period
Correlation analysis between RBP level in urine and renal function test parameters in Table 3

The outcome of MGN patients at the end of follow-up
3.5.1. The outcome of primary MGN patients using eGFR (< 60 ml/min/1.73m 2 ) A prognostic outcome for chronic kidney disease was determined using eGFR (< 60 ml/min/1.73m 2 ); most variables except gender and albumin were preserved following a simple logistic regression as defined in Table 4. However, a  Table 5. For any rise in serum urea in mmol/l, the risk of low eGFR (< 60 ml/min/1.73m 2 ) increases by 3.236.
In contrast, high urinary RBP titre in pg/ml increases the likelihood of a primary outcome (< 60 ml/min/1.73m 2 ) by an odd of 12.987 compared to those with normal urinary RBP titre.

Discussion
The importance of urinary RBP in determining the prognosis of primary MGN compared to secondary MGN was emphasized in this study.
Sixty-nine patients comprising 47 primary and 22 secondary MGN were enrolled in this research. The research emphasized the function of urinary RBP as a prognostic biomarker for chronic kidney disease. It may play a role in the monitoring of those primary MGN patients at risk of ESRD progression. This is in complement with a similar study conducted on the same patients, which denoted the importance of anti-PLA2R and anti-THSD7A in determining the prognosis of primary MGN patients (15).

Role of urinary RBP in monitoring primary MGN patients
A laboratory parameter such as eGFR, known to be used in monitoring MGN patients at risk of progressing to ESRD using CKD classification (16,17) was found to have a significant negative relationship with urine RBP. This implies that lower eGFR is associated with a higher urine RBP titre. Also, UPCr Index used in monitoring primary MGN subjects (especially those presenting with nephrotic syndrome) is significantly correlated with urine RBP. It was also reported that high urinary levels of RBP predicted poor proteinuria outcomes (18). This demonstrated that urinary RBP could be used in monitoring MGN subjects and in the treatment decision in patients with primary MGN.

Urine RBP and prognosis
RBP was discovered to be a reliable biomarker for MGN, predicting those at risk of ESRD (1). In this study, a high titre of urinary RBP was detected more frequently among primary MGN subjects with lower eGFR, higher creatinine and urea levels. Regression analysis conducted to detect factors associated with progression to ESRD in primary MGN subjects demonstrated the persistence of urinary RBP as the main predictor of progression to ESRD. This finding was also supported by a similar study which predicted that urinary RBP could serve as a biomarker for general kidney diseases and not only the proximal convoluted tubule dysfunction (1). Regression analysis in tables 6 and 7 suggested that urinary RBP is associated with achieving remission. Therefore, urinary RBP could serve as an important biomarker for the prognosis of primary MGN. Therefore, higher urinary RBP titre is associated with remission and vice versa.