Abstract

Abstract Background and Aims PXDN is a multifunctional heme peroxidase responsible for the crosslinking of type IV collagen protomers at basement membranes, playing a crucial role in the extracellular matrix dynamics. In renal pathophysiology, increased PXDN expression has been associated with pro-fibrotic and pro-carcinogenic mechanisms and we have shown that PXDN is expressed throughout all kidney compartments since the early stages of nephrogenesis. However, the role of PXDN in kidney health and disease remains largely unknown. In this work we performed a comparative analysis of PXDN expression between renal biopsy samples from IgA nephropathy (IgAN) patients and healthy kidney biopsy samples. IgAN was the selected CKD model due to its straightforward diagnosis by standard protocols for histopathological evaluation of kidney biopsies, and to the fact that the renal outcomes are consistently associated with several key pathologic features, including tubular atrophy and interstitial fibrosis. Correlations between PXDN expression and IgAN disease and kidney function parameters as well as with ECM proteins were also explored. Method Fluorescence immunohistochemistry for PXDN was performed in biopsy samples from IgAN patients (n = 20) and in biopsies from healthy kidney donors (n = 7). Proximal tubules were assessed by anti-CD15 fluorescent staining. Collagen III (COLIII) deposition was investigated in IgAN samples (n = 7) by immunofluorescence. Brightfield images were manually segmented into glomeruli, proximal and distal tubules, and interstitium, using QuPath software, and the fluorescence intensity measurements were obtained with ImageJ/Fiji. Results Results showed that, in comparison with healthy kidney samples, PXDN is significantly decreased in proximal and distal tubules of IgAN patients (P < 0.001 and P < 0.05, respectively) whereas it is increased in IgAN interstitium (P < 0.001). When IgAN samples were stratified by the presence or absence of chronic lesions (glomerular sclerosis and interstitial fibrosis) we observed that in the proximal tubules, PXDN is decreased even in areas without lesions while in the distal tubules the expression is compromised only in the lesioned areas. In the interstitium, PXDN shows a tendency of increase from healthy tissue to IgAN in areas without and with tubulointerstitial fibrosis, presenting a significant increase in the lesioned areas (P < 0.001). Concerning ECM deposition, results showed a significant correlation between the expression of PXDN and COLIII in sclerotic glomeruli (R = 0.8214; P = 0.0341) and in the tubulointerstitial fibrotic areas (R = 0.93; P = 0.0067). Moreover, although no differences were observed in the glomerular expression of PXDN between healthy and IgAN samples, when considering eGFR, a significant increase of PXDN was observed in both non-lesioned and sclerotic glomeruli from IgAN patients with eGFR bellow 60 ml/min/1,73 m2. In the interstitium, the expression of PXDN was also increased in tubulointerstitial fibrotic areas of IgAN patients with eGFR bellow 60 ml/min/1,73 m2, in comparison with the same areas in patients with higher eGFR values. Conclusions Overall, our study shows a differential expression of PXDN in IgAN, with loss in the tubular region that relates with the state of interstitial fibrosis, and gain in the glomerular and interstitial region where it correlates with COLIII compartment-specific deposition and with kidney function as assessed by eGFR. These data support, for the first time, the enrollment of PXDN in the topographic mechanism of IgA-mediated kidney lesion that deserves further investigations. Given that sclerosis and fibrosis are a common pathogenic mechanism of CKD, irrespective of the underlying etiology, we postulate that PXDN may serve as a novel therapeutic target for this disease.

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