Mechanisms of human kidney stone formation

Abstract

The precise mechanisms of kidney stone formation and growth are not completely known, even though human stone disease appears to be one of the oldest diseases known to medicine. With the advent of the new digital endoscope and detailed renal physiological studies performed on well phenotyped stone formers, substantial advances have been made in our knowledge of the pathogenesis of the most common type of stone former, the idiopathic calcium oxalate stone former as well as nine other stone forming groups. The observations from our group on human stone formers and those of others on model systems have suggested four entirely different pathways for kidney stone formation. Calcium oxalate stone growth over sites of Randall's plaque appear to be the primary mode of stone formation for those patients with hypercalciuria. Overgrowths off the ends of Bellini duct plugs have been noted in most stone phenotypes, do they result in a clinical stone? Micro-lith formation does occur within the lumens of dilated inner medullary collecting ducts of cystinuric stone formers and appear to be confined to this space. Lastly, cystinuric stone formers also have numerous small, oval, smooth yellow appearing calyceal stones suggestive of formation in free solution. The scientific basis for each of these four modes of stone formation are reviewed and used to explore novel research opportunities.

Figure 1. Attached stone to site of Randall’s plaque in ICSF patient

Panel a shows an endoscopic view of a calcium oxalate stone (arrow) attached to the tip of a papilla. Several sites of interstitial (Randall’s) plaque (arrowheads) are seen around the attached stone. Note the normal appearance of the papilla. Panel b shows the same papilla after the stone was removed. The papillary surface of that same stone is seen by light microscopy as an inset at the bottom left of this panel. A small site of whitish mineral (marked by asterisk) is clearly visible and was identified as hydroxyapatite while the rest of the stone is calcium oxalate. After performing micro-CT imaging of this stone, these images were aligned and superimposed onto the papilla in order to determine if the small site of hydroxyapatite aligned with a region of Randall’s plaque on the papilla. Indeed, the small apatite region on the stone aligned with a small bleed (see circled area) on a site of interstitial plaque, presumably the site where the stone was attached to the papillum.

Figure 2. Histologic images showing initial sites of Randall’s plaque and its progression

Light microscopy reveals the initial sites of interstitial deposits (panel a, arrows) to be in the basement membranes of the thin loops of Henle at the papilla tip. By transmission electron microscopy, these sites of interstitial deposits are made up of numerous micro-spherulites of alternating lamina of matrix with and without crystals (panels b–e). The individual deposits are as small as 50 nm and grow into multi-layered spheres of alternating light and electron dense rings with the light regions representing crystals and the electron dense sites matrix material. Extensive accumulation of crystalline deposits occur around the loops of Henle (panel f, double arrows) spread into the nearby interstitial space extending to the urothelial lining (panel g) of the urinary space. Disruption of the urothelial layer exposes the site of interstitial deposits to the urine which can trigger overgrowth of mineral and thus stone formation.

Figure 3. Ultrastructural features of attachment site of kidney stone from idiopathic calcium oxalate stone former

Panel a is an endoscopic view of a half millimeter stone seen attached to papilla tip at a site of Randall’s plaque (arrow) while panel b shows the same stone by light microscopy with underlying tissue (arrow) after biopsy. Panel c shows the same stone-tissue complex seen in panel b but after demineralization. Note that the stone separated from the underlying tissue (rectangular box). Some tissue is still stuck to stone (asterisk) while the double arrowheads show a region of cellular debris. The single arrows point to areas on the papilla that still have a urothelial covering, these cells are lost at the stone-tissue junction. Panel d is a high magnification transmission electron micrograph of the tissue attachment site. The region of Randall’s plaque (lower right) is seen covered by a multi-layer, ribbon-like structure, have crystalline and matrix material, which are highlighted in the insert (upper right). A region marked “A” shows small (asterisk) and large crystals (single arrows) embedded in the outer (urine) side of the ribbon.

Figure 4. Calcium oxalate stones grows on sites of interstitial (Randall’s) plaque

The earliest nuclei of plaque appear in the basement membrane of the thin loops of Henle (left most cartoon) as multi-layered spherulites of crystals and matrix material. Subsequently, interstitial plaque genesis (second cartoon, upper row) seems to occur on type 1 collagen in the interstitial space where individual spherulites appear to fuse. With time, islands of matrix and spherulites spread to the basal side of the urothelium and the plaque remains protected from the urine until the urothelium is breached (cartoon 3, upper row). At this point the urine appears to create nucleations that form the base of an overgrowth (cartoons 4 to 8). The urine proteins that form the immediate overlay include Tamm Horsfall protein (THP) and osteopontin (OP) followed by amorphous hydroxyapatite. As the overgrowth region expands there is a conversion of hydroxyapaittie to calcium oxalate (CaOx), the primary mineral of the stone of a ICSF stone patient.

Figure 5. Endoscopic and histologic images from brushite stone formers

Papilla from BR patients (panel a) often showed depressions (arrows) near the papillary tip and flatting, a phenomenon not seen in the CaOx stone formers. Like CaOx patients, the papilla from brushite stone formers possessed sites of Randall’s plaque (arrowheads), though less in amount. In addition, the papilla possessed sites of yellowish crystalline deposit at the openings of ducts of Bellini (asterisk). These ducts were usually enlarged and occasional filled with a crystalline material that protruded from the duct that might serve as a site for stone attachment. Panel b shows crystal deposition in a papillary biopsy stained by the Yasue method. Note crystal deposits in the lumens of individual inner medullary collecting ducts (arrows) and an occasional nearby loop of Henle. The crystal deposits have greatly expanded the lumen of these tubules and induced cell injury to complete cell necrosis in all of these same tubules. A cuff of interstitial inflammation and fibrosis accompanies sites of intra luminal disposition. Panel c shows a third (type 3) pattern of crystal deposits in papillary biopsies of brushite stone formers. Yellowish mineral deposition (single arrows) was found within lumens of medullary collecting ducts just like that described at the opening of ducts of Bellini, except that the type 3 deposits are located in collecting ducts just beneath the urothelium. Sites of type 3 deposits ranged from large areas of crystal deposition in collecting tubules that formed a spoke and wheel-like pattern around the circumference of the papilla to small, single sites of yellowish material in focal regions of a collecting duct lumen. Histologic analysis of the type 3 deposits (panel d) confirms that these sites of crystal deposition are in medullary collecting ducts (asterisk) positioned just beneath the urothelial lining (arrow). The double arrow shows a site of interstitial crystal deposition like that seen in idiopathic CaOx stone formers. Panel’s e and f were obtained from a papillary biopsy collected from a brushite patient here the gross morphology if all papilla was severely injured. Extensive regions of interstitial fibrosis surround the injured collecting ducts (double arrows). Entrapped injured thin loops of Henle and vasa recta (*) are noted in these fields of interstitial fibrosis. A series of crystalline-filled collecting tubules (arrow) are seen embedded in a small ring of fibrotic tissue. An occasional giant cell (arrowheads) was observed near damaged collecting ducts.

Figure 6. High resolution micro-CT images of the BD plugs with overgrowths

Panels a, c, e, and g show a light micrograph of a BD plug with overgrowth region from an apatite, brushite, Ileostomy, and primary hyperparathyroidism stone patient respectively. Panels b, d, f, and h show a corresponding micro-CT image generated from 3D image stacks rendered for surface view and then sized for internal view from each of the plug/overgrowth structure. A dotted line demarks the region of the plug from the overgrowth which would occur at the opening of the duct of Bellini. The mineral forming the plugs from apatite (panel a–b), and primary hyperparathyroid (panels g–h) is entirely apatite. The plug from the brushite patient (panels c–d) is a mixture of brushite, apatite and CaOx while the plug from the ileostomy (panels e–f) is apatite and urates. The overgrowth regions from apatite and primary hyperthyroid patients are primarily concentric layers of apatite, entirely CaOx for ileostomy patients and a mixture of CaOx and and BR for BR patients.

Figure 7. Pelvic stone that possibly started as an overgrowth on a BD plug

This figure shows a long (about 2 cm) and slender renal pelvic stone removed from a patient with primary hyperparathyroidism and viewed by light microscopy and micro-CT. By CT the stone is seen as composed of apatite with a round inclusion region (denoted by arrow and ring) at one side of the stone. Possibly this inclusion region was the original plug because its layers of HA are like those of plugs in being nodular vs the concentric rings of most stones as is seen here.

Figure 8. Cortical Interstitial fibrosis, tubular atrophy and glomerulosclerorsis in brushite (black circles), apatite (open circles) and ICSF (black triangles)

Left panel. After scoring cortical biopsies from BR, apatite and ICSF stone formers for interstitial fibrosis, tubular atrophy and glomerular injury, the highest score for interstitial or tubular atrophy was plotted on the x-axis and glomerular injury on the y axis for each phenotype. Out of 30 cases plotted, only 2 ICSF had interstitial changes above a score of 1, and none had significant glomerular disease with a glomerular score above 1, while 4/11 apatite and 12/25 BRSF had significant cortical injury. Right Panel’s a–c shows the range of interstitial (arrows), tubular and glomerular (G) changes seen by light microscopy in the apatite and brushite patients. Panel a shows the cortex of an apatite patient with mild interstitial fibrosis and tubular atrophy (score = 1) and no glomerulosclerorsis while panel b shows an apatite patient with moderate interstitial fibrosis, tubular atrophy and glomerulosclerorsis (score =2). Panel c shows the cortex of a brushite patient with severe interstitial fibrosis, tubular atrophy and glomerulosclerorsis (score =3).

Figure 9. Endoscopic unroofing of an IMCD ductal stone in cystinuric stone former

Micro-liths of cystine are present at the distal ends of IMCD and are easily seen at the time of percutaneous nephrolithotomy to lie under the urothelium at a site marked by a dark shadow (arrow) on a dilated duct (panel a). When the IMCD is unroofed with a laser (panel b), an unattached, round tiny ‘stone’ is exposed (double arrow) within dilated IMCD at easily flows out of the IMCD lumen.

Figure 10. Cystine stones may be example of kidney stones formed in ‘free solution’

While numerous unattached stones are seen at the time stone removal by percutaneous nephrolithotomy or ureteroscopy only the stones from cystinuric patients have consistent characteristics of a stone formed in ‘free solution’ (panel a). The cystine stones look like ‘Easter eggs” in that they are completely smooth, oval in shape, have a homogenous yellow coloration and are freely floating in a renal calyx. They are easily grasped and removed (panel b).