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Sources of Funding: 2016 Urology Care Foundation Research Scholar Award

Introduction

Hydroxyapatite (HA) is a common constituent of most idiopathic calcium oxalate (CaOx) stones. It is frequently found at the CaOx crystal nucleation site deep within a stone and acts as suitable nucleator of CaOx in vitro. Although most crystal deposits within tissue produce inflammation (brushite and calcium oxalate), renal interstitial HA deposits in idiopathic calcium oxalate stone formers do not produce inflammation but instead accumulate within the interstitium as Randall&[prime]s plaque. To further explore this lack of response, we investigated the effect of oxalate and hydroxyapatite to differentiate primary human monocyte to macrophages.

Methods

In hexaplet, primary human monocytes were exposed to 0.5, 1.0, 2.0, and 3.0mM of CaOx and HA crystals. GM-CSF and M-CSF were included as positive controls for M1 and M2 macrophage differentiation respectively. At day 6, macrophages were washed 3 times with PBS to remove any undifferenced monocytes, and 200ng of LPS from S. enterica serotype Minnesota Re595 was added to half of each treatment group. After 4hrs, total RNA was collected, and cytokine expression was analyzed by qPCR.

Results

By day 3, monocytes exposed to CaOx and HA displayed macrophage morphology. At day 6, CaOx, HA, GM-CSF, and M-CSF displayed complete macrophage morphology (Figure). CaOx differentiates macrophages displayed similar inflammatory cytokine (TNFα, IL-6, and IL-1β) expression as GM-CSF; both displayed several hundred-fold greater inflammatory cytokine expression than HA and M-CSF differentiated macrophages.

Conclusions

In our immunological model for stone formation, CaOx and HA display competing immunological effects. HA induces tissue healing (M2)-like macrophages whereas CaOx induces inflammatory (M1)-like macrophages. In the context of Randall's plaque, we speculate that luminal CaOx crystals induce this potent M1 inflammatory response in monocytes, which can trigger renal epithelial cell production of HA. This HA (Randall&[prime]s plaque) induces monocyte differentiation into tissue healing (M2) macrophages, blocking further M1 inflammation. This may explain the lack of significant papillary inflammation in the pathogenesis of CaOx stone formation.

Funding

2016 Urology Care Foundation Research Scholar Award