Background/Purpose: Systemic Lupus Erythematosus (SLE) is a chronic autoimmune disease predominantly affecting women of childbearing age. Roughly 50% of SLE patients develop Lupus Nephritis (LN), a serious kidney disease associated with higher morbidity and mortality. Current urinalysis markers, such as the urine protein to urine creatinine ratio (UPr:Ucr), lack specificity for glomerular damage, necessitating repeated kidney biopsies to monitor disease progression. This highlights the need for sensitive, non-invasive diagnostic and prognostic urine biomarkers.

We propose that glycosylation changes in the endothelial glycocalyx precede irreversible damage detectable by current urine markers. Our previous studies identified elevated glycosphingolipids and an altered N-glycome as discriminative LN markers. We hypothesize that urinary glycosylation changes, detected by matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI), can differentiate LN from SLE and healthy controls, offering insights into glomerular damage and potentially predicting treatment response.

Methods: Utilizing mass spectrometry methods pioneered by our lab, we analyzed 405 de-identified urine samples from LN patients and controls for N-glycan composition. Each sample underwent treatment with PNGase F to remove N-glycans from glycoproteins and was then analyzed using MALDI-MSI after application of a MALDI matrix solution. Data processing was conducted using SCIMAX Mass Spectrometry instrumentation and SCILS lab software. N-glycan spectra were annotated by matching glycan peak m/z values to an in-house database, utilizing accurate mass determinations from MALDI-QTOF instrumentation. Renal biopsy samples were also analyzed using the same MALDI workflow to correlate urinary glycomic signatures with renal histology.

Results: In our initial analysis of 405 urine samples, we identified a total of 81 unique N-glycans, with 45 showing statistical significance in a one-way ANOVA (p-value of 5 x 10^-5). Sparse partial least squares discriminant analysis revealed a clear distinction between LN and control (SLE/HC) samples (Figure 1). Analysis of total N-glycans showed that LN urine had elevated levels of high mannose and sialylated N-glycans, while core-fucosylated N-glycans were more prevalent in HC and SLE samples, possibly indicating a “healthy” glycocalyx.

Analysis of LN renal biopsy samples using the 81 unique N-glycans identified in LN urine samples revealed correlations between urinary glycomic signatures and renal histology. Specific N-glycan intensities were linked to kidney structures, with clear distinctions such as proximal convoluted tubule-specific glycans like m/z 2669.96 (Figure 2) and glomerular-specific glycans like m/z 2539.90 (Figure 3).

Conclusion: This study demonstrates the potential of urinary N-glycans as biomarkers of disease. Importantly, these markers may offer advantages over current urinalysis methods by reflecting histological changes in the kidney, indicating promising avenues for future research and clinical application.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/glycosylation-changes-as-non-invasive-biomarkers-for-lupus-nephritis-detection-and-prognosis/