Immunization with Pentraxin3 prevents transition from subclinical to clinical lupus nephritis in lupus-prone mice: Insights from renal ultrastructural findings

Highlights

• Immunization with pentraxin3 (PTX3) protects against lupus nephritis development.

• PTX3 colocalizes with nuclear material in lupus glomeruli.

• PTX3 is mostly localized inside glomerular electron-dense deposits.

• PTX3 is a novel driving autoantigen that may sustain renal inflammation.

Abstract

Background

Pentraxin3 (PTX3) is an emerging player in lupus nephritis (LN). Anti-PTX3 antibodies showed to delay LN occurrence in vivo.

Aim

To evaluate renal changes following immunization with PTX3 in a murine model of LN.

Materials and methods

Twenty-two lupus-prone New Zealand Black/White (NZB/W)F1 mice were divided into two groups (n = 11) and subcutaneously injected with human recombinant (hr)PTX3 100 μg or phosphate buffer saline (PBS) 200 μl, three times 3 weeks apart, starting before development of proteinuria. Five mice from each group were scheduled for sacrifice at week 22 and 6 from each group at week 29. Renal lesions included electron-dense deposits (EDD), glomerular deposition of IgG, complement and PTX3 as markers of renal inflammation. They were evaluated by immunofluorescence (IF), confocal and immunoelectron microscopy (IEM). Validated semiquantitative scores were used when available to score renal lesions. Chi-squared test with Fisher exact test was used for comparison.

Results

Nineteen out of 22 mice were sacrificed as scheduled. Only hrPTX3-immunized mice developed anti-PTX3 antibodies. Compared to PBS-injected mice, they displayed a dramatic decrease in glomerular deposits of IgG, C1q and PTX3, as well as in the amount of EDD (p = 0.006) and podocyte effacement (p = 0.043). Importantly, PTX3 was pinpointed inside the EDD and co-localized with nuclear material.

Conclusions

Immunization with PTX3 prevented progression from the preclinical to the clinical stage of LN, inciting anti-PTX3 antibodies and preventing renal PTX3 deposition. PTX3 is a novel component of EDD, submitting it as one initiating autoantigen in LN and as potential target for early treatment.

Introduction

Lupus glomerulonephritis (LN) is one the most severe and common manifestations of systemic lupus erythematosus (SLE), affecting up to 50% of patients during their disease course and leading to considerable morbidity and mortality if not properly treated [1,2]. In fact, the rate of renal response following initial treatment is low and the proportion of patients maintaining a long-lasting response is lower than 50% [3], with a remarkable 40% of patients still developing some degree of renal impairment in the long term [4], culminating in end stage renal disease in 10% of the cases [5]. These still unsatisfactory results may reflect the incomplete understanding of the underlying pathogenic mechanisms driving LN occurrence.

The interplay of pentraxin 3 (PTX3) and anti-PTX3 antibodies has emerged in the last years as a likely component of LN development [[6], [7], [8], [9]]. PTX3 has long been described as an evolutionary conserved molecule responsible for a number of functions in the human organism, especially antimicrobial and homeostatic [10,11]. PTX3 is an acute phase protein involved in opsonization of invading pathogens and/or apoptotic cells with an overall anti-inflammatory activity under physiological conditions. However, in case of extensive tissue damage, e.g. ischemia-reperfusion injury, or in presence of an exaggerated burden of exposed antigens, PTX3 may be shifted towards a pro-inflammatory phenotype resulting in an uncontrolled complement activation and inflammation [[12], [13], [14]]. Accordingly, PTX3 deposition was observed in kidneys of patients with LN where it correlated with the extent of fibrosis and proteinuria [6], while anti-PTX3 antibodies were inversely associated with LN occurrence in both patients and lupus murine models [[6], [7], [8], [9]]. The underlying mechanisms of protection against renal inflammation may be partially due to anti-PTX3 antibodies mostly belonging to IgG4 subclass [9], yet other still unravelled mechanisms are possible.

Descending from the clinical benefit provided by immunization with human recombinant (hr)PTX3 in lupus-prone mice [9], in the current study we aim at assessing the ultrastructural changes induced in mice kidneys upon immunization with hrPTX3, in order to draw a more complete picture of the effects of anti-PTX3 immunity and to explore the effects of an early antigen-targeted treatment when a clinically evident LN is not yet established.