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Observations on the Ultrastructure and Genesis of Urinary Calculi
Vol.117, March Printed in U.S.A.
THE JOURNAL OF UROLOGY
Copyright © 1977 by The Williams & Wilkins Co.
OBSERVATIONS ON THE ULTRASTRUCTURE AND GENESIS OF URINARY CALCULI R.
s.
MALEK*
AND
w.
H. BOYCE
From the Department of Urology, Mayo Clinic and Mayo Foundation, Rochester, Minnesota, and the Division of Urology, Bowman Gray School of Medicine, Winston-Salem, North Carolina
ABSTRACT
Conventional and scanning electron microscopy of calcigerous renal calculi discloses typical concentric laminations, radial striations and microspherules. Random axial distribution of oxalate crystals and their coating by electron-dense matrix fibers with a definite parallel orientation and cross-linkages are evident. The biochemical relationship of uromucoid to matrix substance A is described. It is suggested that renal sialidase may convert the urinary uromucoid to matrix substance A, whose apatitecovered fibers may be responsible for epitaxial nucleation of some crystal systems. Our studies indicate that the intimate apatite-matrix relationship occurs in the human nephron, probably as an intracellular phenomenon. Subsequent extrusion of these mineralized complexes into the lumen of the nephron (intranephronic calculosis) may, in some instances, represent the initial microanatomic stage of renal calculogenesis. In the Western Hemisphere human urinary concretions are predominantly of renal origin and manifest a remarkably characteristic internal architecture despite varied appearance. Their basic morphology may be described in various combinations of concentric laminations (with or without frond formation), radial striations and spherules. 1 No natural concretion is ever formed as a sedimentary mass of crystals. Crystals and matrix are invariably present as fundamental components of each stone and are intimately associated morphologically. Much controversy surrounds the relative importance and mechanisms of the processes by which crystalmatrix interrelations develop.'· 2 In an attempt to clarify some of the controversial issues an ultrastructural demonstration of the organization of calculus matrix and crystals together with a biochemical description of the matrix, its possible precursor and intrarenal development of a crystal-matrix relationship (intranephronic calculosis) as a potential source of renal lithiasis is presented.
interlaminar spaces. Other fibers form radial striations characterized by right angle orientation of the fibers with end-to-side linkages. They always have their terminals at junctures with concentric laminations (fig. 5). At higher magnifications the individual fibers appear to have a ribbon-like structure, with a remarkably constant width-to-breadth ratio of 1 to 5.5 and cross-linkages of unknown periodicity (fig. 6). In contrast to the highly organized structure of the macrospherule the remaining heavily mineralized calculous mass consists of randomly distributed oxalate crystals, with small
ULTRASTRUCTURAL OBSERVATIONS
Conventional or scanning electron microscopy was used for 28 calcium oxalate or phosphate calculi obtained from uninfected idiopathic stone-forming patients. The technical methodologic details have been described elsewhere. 3 Basically, such calculi consist of 2 parts: 1) a large pale mass of surface crystals and 2) a dark, mechanically removable macrospherule of calcium phosphate and oxalate, measuring approximately 1 mm. in diameter (fig. 1). The finely nodular outer surface of the macrospherule is covered by densely mineralized matrix and its cut surface contains areas of amorphous, poorly mineralized gelatinous matrix (fig. 2). Within the gelatinous matrix collections of minute laminated or vacuolated spheres are observed regularly (fig. 3). Currently, their nature and role are speculative. The highly organized fibrous matrix consists of parallel bundles of fibers, some of which form concentric laminations. The individual fibers vary in length and change their orientation with that of the lamination (fig. 4). Amorphous matrix fills the Accepted for publication June 25, 1976. Read at annual meeting of American Urological Association, Las Vegas, Nevada, May 16-20, 1976. * Requests for reprints: Mayo Clinic, 200 First St., S.W., Rochester, Minnesota 55901. 336
Fm. 1. A, calcium oxalate calculus (3 by 5 mm.) with large pale mass of surface crystals and dark macrospherule. B, macrospherule mechanically removed from calculus.
337
ULTRASTRUCTDRE AND GENESIS OF CALCULI
FIG. 2. A, scanning electron micrograph of nodule from surface of macrospherule. Reduced from like cut surface of macrospherule. Reduced from x500.
FIG. 3. Electron micrograph of 2 collections of minute laminated or vacuolated spherules surrounded by electron-dense membrane in amorphous matrix. Reduced from x27,000.
ones budding off the surfaces of large ones (fig. 7). Closer inspection discloses parallel electron-dense matrix fibers, with cross-linkages, sweeping over and interlocking the crystal facies (fig. 8). Another generation of small spheres, the microspherules, is found frequently among oxalate crystals (fig. 9). They are far more abundant in calcium phosphate calculi (fig. 10) and bear a remarkable resemblance to Carr's intralymphatic microliths (fig. 11, F). BIOCHEMICAL CONSIDERATIONS
The matrix is a mucoprotein. It exists in fibrous and amorphous gel-like forms and on a weight-to-weight basis accounts for 2.5 per cent (in calcigerous calculi) to 62 per cent (in matrix calculi) of the calculous mass. Its basic chemical composition is similar to that of most of the human mucoids: protein 65 per cent, carbohydrate 14 per cent, bound water 10 per cent and bound inorganic ash 12 per cent. However, it differs from bone collagen in that it contains neither hydroxyproline nor elastin (less than 2 per cent praline). 1 • •. 5 However, the absence of fibrous proteins does not preclude hydroxyapatite epitaxy by the matrix since its free amino groups are capable of performing such a task. 1 Immunologic techniques have provided practically all the current information about the matrix components. Substance A, which is the most potent antigenic component of all urinary calculi, forms 85 per cent of the matrix mass. The other com-
X 750.
B, scanning electron micrograph of gel-
ponents of matrix include serum albumin, 1 or 2 alpha-globulins and, occasionally, gamma-globulins and uromucoid. 1 Matrix substance A is apparently of renal origin because it is detectable immunologically in renal tissue of patients with renal calculi, in some diseased kidneys with microscopic calcification and in alkaline secretions of the alimentary tract. It is not detectable in normal kidneys. 1 ' • The same substance is present in large quanhties (23 mg. per 24 hours) in the urine of active stone formers (R-S 1 fraction). Its concentration is much decreased in the urine of occasional stone formers and it is absent in normal urine, 1 • 6 The over-all composition of matrix from urinary calculi of various crystalline composition is sufficiently simil;r to suggest a common precursor. No less than 7 such precursors, including uromucoid, have received attention. 7 Uromucoid forms the nondialyzable insoluble fraction (R-1) of the urinary macromolecules, whose major component is Tamm and Horsfall virusinhibitory mucoprotein. Immunologic techniques have identified the renal cortex and medulla as its only source. 8 It is found in all normal human urine and is present in increased concentration in the urine of stone formers and in patients with inflammatory diseases of the urinary passages. s, 8 Conversely, racially pure blacks, who practically never have stones, have no uromucoid in their urine. 8 The composition of uromucoid closely resembles that of stone matrix, the main difference being the relative quantities of glucide monomers. 5 Sialic acid, usually a terminal unit of the carbohydrate fraction, is often present in uromucoid but is consistently absent from stone matrix. 8 Because sialidase (N-acetyl neuraminidase) is one of the regularly present renal enzyme systems, this observation suggests that the first step in conversion of mucosubstances to mineralizable matrix may be the removal of the neuraminic group from the glucide moiety. Crystals of organic and inorganic composition form the major portion of the common urinary calculi on a weight-to-weight basis. Invariably, the fibrous matrix of even the purest calculi of any crystalline composition is covered by crystals that are too small to be resolved in the native state by the current scanning electron microscopic techniques. They are presumed to be calcium phosphate, as apatite, on the basis of examination by various other techniques-microradiography, polarizing microscopy and chemical analysis. 1 Calcium oxalate (monohydrate and dihydrate) crystals appear in greatest concentrations on the apatite-covered fibers but fill the interlaminar matrix in variable degrees, depending on the extent of mineralization of the particular stone. Uric acid and magnesium ammonium phosphate crystals, on the other hand, have a predilection only for the interlaminar amorphous matrix. 1
338
MALEK AND BOYCE
FIG. 4. Electron micrograph of concentric laminations of macrospherule. A, fibrous (dark) and interlaminar (light) matrix. Reduced from x 5 700. '
B, change in fiber orientation with directional change in concentric lamination. Reduced from x 13,800.
FIG. 5. Electron micrograph of juncture of radial striations and concentric laminations. Reduced from x27,000.
FIG. 6. Electron micrograph of matrix fibers showing ribbon-like fibril structure (inset). Reduced from x82,500.
INTRARENAL CALCULOGENESIS
The intimate relationship of calcium phosphate to fibrous matrix is a universal phenomenon and is demonstrable in all natural human urinary concretions regardless of size. This occurs in the human nephron principally in the distal tubule, probably as high as the proximal tubule, and possibly as an intracellular phenomenon. 9 Laminated spherules of hydroxyapatite-mucosubstance complexes (periodic acid, Schiff positive) may be associated with disruptions of the mitochondria of the renal tubular cells under a variety of circumstances in animals and in man (fig. 11, A and B). 9 Extrusion of these spherules into the lumen of the nephron and further mineralization may represent the initial stage of renal calculogenesis (fig. 11, C and D). Our studies of fresh renal biopsy specimens obtained from 100 stone-forming and non-stone-forming patients indicate that this phenomenon of intranephronic calculosis is indeed demonstrable in all idiopathic stone formers. 9 Its intensity is directly proportional to the activity of the stone disease and its nature is fundamentally not different from Randall's submucosal plaques or Carr's intralymphatic microliths (fig. 11, E and F). 9 Intranephronic calculosis usually is not observed in the kid-
neys of individuals with struvite, uric acid or cystine calculi. In such patients with pathologic deviation of normal biologic mineralization, epitaxial nucleation of hydroxyapatite by the matrix may initially occur in the abnormal pelviocaliceal urine. 9 COMMENT
Urinary calculogenesis is not a singular, simple or direct process but the result of multiple, complex and interrelated phenomena. Despite extensive research the exact mode of stone formation is still speculative. The consistent presence of an architecturally well organized matrix and its intimate association with hydroxyapatite are demonstrable in practically all visible and detectable phases of natural human calculogenesis. The importance of the matrix-apatite relationship is further emphasized by recent in vitro studies demonstrating the ability of apatite crystals to induce epitaxial growth of calcium oxalate monohydrate crystals. 10 • These observations on the various phases of calculogenesis indicate that perhaps most of the more common renal calculi may be initially conceived as crystal-matrix organizations within the nephron. Exactly how and, more importantly, under
ULTRASTRUCTURE AND GENESIS OF CALCULI
839
FIG. 7. Scanning electron micrograph of surface crystals of calculus seen in figure 1, A. A, note random distribution of crystals. Reduced from x 150. B, small calcium oxalate crystal budding off surface of large crystal. Reduced from x750.
FIG. 8. Scanning electron micrograph of surface crystals of calculus seen in figure 1, A. A, closer inspection shows matrix fibers covering and interlocking crystal facies. Reduced from x 750. B, note distinct parallelism of matrix fibers with cross-linkages covering crystal facies. Reduced from x375.
FIG. 9. Scanning electron micrograph of surface structure of calculus seen in figure 1, A shows microspherule (center). Reduced from x75.
FIG. 10. Scanning electron micrograph of surface structure of calcium phosphate calculus shows abundance of microspherules. Reduced from x 1,500.
FIG. 11. Intracellula r spherules: A , collection of large positive spherules of mucosubstance in renal tubular cells of actively stone-forming patient. PAS stain. B , collection of laminated calcified spherules in renal tubular cells, distinctly visible near lumina of tubules. Alizarin red stain. Intranephronic calculi within lumen of human nephron: C, laminated mucosubstance microlith. PAS, reduced from x 160. D, calcification of same laminated microlith. Alizarin red, reduced from x 155. Randall's plaque and Carr's bodies: E, submucosal collection of Randall's plaques (arrow) near t ip ofrenal papilla. Alizarin red stain. F, scanning electron micrograph of Carr's intralymphatic microlith. Reduced from x 50.
340
341
ULTRASTRUCTURE AND GENESIS OF CALCULI
what this union takes place within the tubular cel.i or in the abnormal pelviocaliceal urine can only be elucidated by further research. Mr. James W. Willard assisted in these studies. Figures 1 to 10 reprinted with permission. 3 Figure 11, Falso reprinted with permission from Carr, R. J.: Aetiology of renal calculi: microradiographic studies. In: Renal Stone Research Symposium. Edited by A. Hodgkinson and B. E. C. Nordin. London: J. & A. Churchill, Ltd., pp. 123-132, 1969.
7. 8. 9.
10.
kinson and B. E. C. Nordin. London: J. & A. Churchill, Ltd., pp. 181-189, 1969. Boyce, W. H. and King, J. S., Jr.: Present concepts concerning the origin of matrix and stones. Ann. N.Y. Acad. Sci., 104: 563, 1963. Keutel, H.J., King, J. S., Jr. and Boyce, W. H.: Further studies of uromucoid in normal and stone urine. Urol. Int., 17: 324, 1964. Malek, R. S. and Boyce, W. H.: Intranephronic calculosis: its significance and relationship to matrix in nephrolithiasis. J. Urol., 109: 551, 1973. Meyer, J. L., Bergert, J. H. and Smith, L. H.: Epitaxial relationships in urolithiasis: the calcium oxalate monohydrate-hydroxyapatite system. Clin. Sci. Mo!. Med., 49: 369, 1975.
REFERENCES
1. Boyce, W. H.: Organic matrix of human urinary concretions. Amer.
J. Med., 45: 673, 1968. 2. Vermeulen, C. W. and Lyon, E. S.: Mechanisms of genesis and growth of calculi. Amer. J. Med., 45: 684, 1968. 3. Boyce, W. H.: Some observations on the ultrastructure of "idiopathic" human renal calculi. In: Urolithiasis: Physical Aspects. Edited by B. Finlayson, L. L. Hench and L. H. Smith. Washington, D.C.: National Academy of Sciences, pp. 97-114, 1972. 4. King, J. S., Jr. and Boyce, W. H.: Amino acid and carbohydrate composition of the mucoprotein matrix in various calculi. Proc. Soc. Exp. Biol. Med., 95: 183, 1957. 5. King, J. S., Jr. and Boyce, W. H.: Analysis of renal calculous matrix compared with some other matrix materials and with uromucoid. Arch. Biochem. Biophys., 82: 455, 1959. 6. Boyce, W. H.: Macromolecular components of kidney calculi in urine. In: Renal Stone Research Symposium. Edited by A. Hodg-
COMMENT This report makes us mindful that stone research has gone a full cycle. In the 1950s Butt advanced the notion of protective colloid in the treatment of urinary stone patients, and Boyce and associates presented their pioneer work on characterizing stone matrix. During the 1960s work on the crystalline aspects of urolithiasis became predominant and work on macromolecules as they relate to urolithiasis was largely neglected except by a few workers. Now with a better understanding of the physical chemistry of crystals and electrolyte solutions there is a resurgence of interest in the relation of urinary macromolecules to urolithiasis. It is very much to be desired that work in this area will now advance pari passu with our increasing understanding of the crystalline features of stones. Birdwell Finlayson University of Florida College of Medicine Gainesville, Florida
THE JOURNAL OF UROLOGY
Copyright © 1977 by The Williams & Wilkins Co.
OBSERVATIONS ON THE ULTRASTRUCTURE AND GENESIS OF URINARY CALCULI R.
s.
MALEK*
AND
w.
H. BOYCE
From the Department of Urology, Mayo Clinic and Mayo Foundation, Rochester, Minnesota, and the Division of Urology, Bowman Gray School of Medicine, Winston-Salem, North Carolina
ABSTRACT
Conventional and scanning electron microscopy of calcigerous renal calculi discloses typical concentric laminations, radial striations and microspherules. Random axial distribution of oxalate crystals and their coating by electron-dense matrix fibers with a definite parallel orientation and cross-linkages are evident. The biochemical relationship of uromucoid to matrix substance A is described. It is suggested that renal sialidase may convert the urinary uromucoid to matrix substance A, whose apatitecovered fibers may be responsible for epitaxial nucleation of some crystal systems. Our studies indicate that the intimate apatite-matrix relationship occurs in the human nephron, probably as an intracellular phenomenon. Subsequent extrusion of these mineralized complexes into the lumen of the nephron (intranephronic calculosis) may, in some instances, represent the initial microanatomic stage of renal calculogenesis. In the Western Hemisphere human urinary concretions are predominantly of renal origin and manifest a remarkably characteristic internal architecture despite varied appearance. Their basic morphology may be described in various combinations of concentric laminations (with or without frond formation), radial striations and spherules. 1 No natural concretion is ever formed as a sedimentary mass of crystals. Crystals and matrix are invariably present as fundamental components of each stone and are intimately associated morphologically. Much controversy surrounds the relative importance and mechanisms of the processes by which crystalmatrix interrelations develop.'· 2 In an attempt to clarify some of the controversial issues an ultrastructural demonstration of the organization of calculus matrix and crystals together with a biochemical description of the matrix, its possible precursor and intrarenal development of a crystal-matrix relationship (intranephronic calculosis) as a potential source of renal lithiasis is presented.
interlaminar spaces. Other fibers form radial striations characterized by right angle orientation of the fibers with end-to-side linkages. They always have their terminals at junctures with concentric laminations (fig. 5). At higher magnifications the individual fibers appear to have a ribbon-like structure, with a remarkably constant width-to-breadth ratio of 1 to 5.5 and cross-linkages of unknown periodicity (fig. 6). In contrast to the highly organized structure of the macrospherule the remaining heavily mineralized calculous mass consists of randomly distributed oxalate crystals, with small
ULTRASTRUCTURAL OBSERVATIONS
Conventional or scanning electron microscopy was used for 28 calcium oxalate or phosphate calculi obtained from uninfected idiopathic stone-forming patients. The technical methodologic details have been described elsewhere. 3 Basically, such calculi consist of 2 parts: 1) a large pale mass of surface crystals and 2) a dark, mechanically removable macrospherule of calcium phosphate and oxalate, measuring approximately 1 mm. in diameter (fig. 1). The finely nodular outer surface of the macrospherule is covered by densely mineralized matrix and its cut surface contains areas of amorphous, poorly mineralized gelatinous matrix (fig. 2). Within the gelatinous matrix collections of minute laminated or vacuolated spheres are observed regularly (fig. 3). Currently, their nature and role are speculative. The highly organized fibrous matrix consists of parallel bundles of fibers, some of which form concentric laminations. The individual fibers vary in length and change their orientation with that of the lamination (fig. 4). Amorphous matrix fills the Accepted for publication June 25, 1976. Read at annual meeting of American Urological Association, Las Vegas, Nevada, May 16-20, 1976. * Requests for reprints: Mayo Clinic, 200 First St., S.W., Rochester, Minnesota 55901. 336
Fm. 1. A, calcium oxalate calculus (3 by 5 mm.) with large pale mass of surface crystals and dark macrospherule. B, macrospherule mechanically removed from calculus.
337
ULTRASTRUCTDRE AND GENESIS OF CALCULI
FIG. 2. A, scanning electron micrograph of nodule from surface of macrospherule. Reduced from like cut surface of macrospherule. Reduced from x500.
FIG. 3. Electron micrograph of 2 collections of minute laminated or vacuolated spherules surrounded by electron-dense membrane in amorphous matrix. Reduced from x27,000.
ones budding off the surfaces of large ones (fig. 7). Closer inspection discloses parallel electron-dense matrix fibers, with cross-linkages, sweeping over and interlocking the crystal facies (fig. 8). Another generation of small spheres, the microspherules, is found frequently among oxalate crystals (fig. 9). They are far more abundant in calcium phosphate calculi (fig. 10) and bear a remarkable resemblance to Carr's intralymphatic microliths (fig. 11, F). BIOCHEMICAL CONSIDERATIONS
The matrix is a mucoprotein. It exists in fibrous and amorphous gel-like forms and on a weight-to-weight basis accounts for 2.5 per cent (in calcigerous calculi) to 62 per cent (in matrix calculi) of the calculous mass. Its basic chemical composition is similar to that of most of the human mucoids: protein 65 per cent, carbohydrate 14 per cent, bound water 10 per cent and bound inorganic ash 12 per cent. However, it differs from bone collagen in that it contains neither hydroxyproline nor elastin (less than 2 per cent praline). 1 • •. 5 However, the absence of fibrous proteins does not preclude hydroxyapatite epitaxy by the matrix since its free amino groups are capable of performing such a task. 1 Immunologic techniques have provided practically all the current information about the matrix components. Substance A, which is the most potent antigenic component of all urinary calculi, forms 85 per cent of the matrix mass. The other com-
X 750.
B, scanning electron micrograph of gel-
ponents of matrix include serum albumin, 1 or 2 alpha-globulins and, occasionally, gamma-globulins and uromucoid. 1 Matrix substance A is apparently of renal origin because it is detectable immunologically in renal tissue of patients with renal calculi, in some diseased kidneys with microscopic calcification and in alkaline secretions of the alimentary tract. It is not detectable in normal kidneys. 1 ' • The same substance is present in large quanhties (23 mg. per 24 hours) in the urine of active stone formers (R-S 1 fraction). Its concentration is much decreased in the urine of occasional stone formers and it is absent in normal urine, 1 • 6 The over-all composition of matrix from urinary calculi of various crystalline composition is sufficiently simil;r to suggest a common precursor. No less than 7 such precursors, including uromucoid, have received attention. 7 Uromucoid forms the nondialyzable insoluble fraction (R-1) of the urinary macromolecules, whose major component is Tamm and Horsfall virusinhibitory mucoprotein. Immunologic techniques have identified the renal cortex and medulla as its only source. 8 It is found in all normal human urine and is present in increased concentration in the urine of stone formers and in patients with inflammatory diseases of the urinary passages. s, 8 Conversely, racially pure blacks, who practically never have stones, have no uromucoid in their urine. 8 The composition of uromucoid closely resembles that of stone matrix, the main difference being the relative quantities of glucide monomers. 5 Sialic acid, usually a terminal unit of the carbohydrate fraction, is often present in uromucoid but is consistently absent from stone matrix. 8 Because sialidase (N-acetyl neuraminidase) is one of the regularly present renal enzyme systems, this observation suggests that the first step in conversion of mucosubstances to mineralizable matrix may be the removal of the neuraminic group from the glucide moiety. Crystals of organic and inorganic composition form the major portion of the common urinary calculi on a weight-to-weight basis. Invariably, the fibrous matrix of even the purest calculi of any crystalline composition is covered by crystals that are too small to be resolved in the native state by the current scanning electron microscopic techniques. They are presumed to be calcium phosphate, as apatite, on the basis of examination by various other techniques-microradiography, polarizing microscopy and chemical analysis. 1 Calcium oxalate (monohydrate and dihydrate) crystals appear in greatest concentrations on the apatite-covered fibers but fill the interlaminar matrix in variable degrees, depending on the extent of mineralization of the particular stone. Uric acid and magnesium ammonium phosphate crystals, on the other hand, have a predilection only for the interlaminar amorphous matrix. 1
338
MALEK AND BOYCE
FIG. 4. Electron micrograph of concentric laminations of macrospherule. A, fibrous (dark) and interlaminar (light) matrix. Reduced from x 5 700. '
B, change in fiber orientation with directional change in concentric lamination. Reduced from x 13,800.
FIG. 5. Electron micrograph of juncture of radial striations and concentric laminations. Reduced from x27,000.
FIG. 6. Electron micrograph of matrix fibers showing ribbon-like fibril structure (inset). Reduced from x82,500.
INTRARENAL CALCULOGENESIS
The intimate relationship of calcium phosphate to fibrous matrix is a universal phenomenon and is demonstrable in all natural human urinary concretions regardless of size. This occurs in the human nephron principally in the distal tubule, probably as high as the proximal tubule, and possibly as an intracellular phenomenon. 9 Laminated spherules of hydroxyapatite-mucosubstance complexes (periodic acid, Schiff positive) may be associated with disruptions of the mitochondria of the renal tubular cells under a variety of circumstances in animals and in man (fig. 11, A and B). 9 Extrusion of these spherules into the lumen of the nephron and further mineralization may represent the initial stage of renal calculogenesis (fig. 11, C and D). Our studies of fresh renal biopsy specimens obtained from 100 stone-forming and non-stone-forming patients indicate that this phenomenon of intranephronic calculosis is indeed demonstrable in all idiopathic stone formers. 9 Its intensity is directly proportional to the activity of the stone disease and its nature is fundamentally not different from Randall's submucosal plaques or Carr's intralymphatic microliths (fig. 11, E and F). 9 Intranephronic calculosis usually is not observed in the kid-
neys of individuals with struvite, uric acid or cystine calculi. In such patients with pathologic deviation of normal biologic mineralization, epitaxial nucleation of hydroxyapatite by the matrix may initially occur in the abnormal pelviocaliceal urine. 9 COMMENT
Urinary calculogenesis is not a singular, simple or direct process but the result of multiple, complex and interrelated phenomena. Despite extensive research the exact mode of stone formation is still speculative. The consistent presence of an architecturally well organized matrix and its intimate association with hydroxyapatite are demonstrable in practically all visible and detectable phases of natural human calculogenesis. The importance of the matrix-apatite relationship is further emphasized by recent in vitro studies demonstrating the ability of apatite crystals to induce epitaxial growth of calcium oxalate monohydrate crystals. 10 • These observations on the various phases of calculogenesis indicate that perhaps most of the more common renal calculi may be initially conceived as crystal-matrix organizations within the nephron. Exactly how and, more importantly, under
ULTRASTRUCTURE AND GENESIS OF CALCULI
839
FIG. 7. Scanning electron micrograph of surface crystals of calculus seen in figure 1, A. A, note random distribution of crystals. Reduced from x 150. B, small calcium oxalate crystal budding off surface of large crystal. Reduced from x750.
FIG. 8. Scanning electron micrograph of surface crystals of calculus seen in figure 1, A. A, closer inspection shows matrix fibers covering and interlocking crystal facies. Reduced from x 750. B, note distinct parallelism of matrix fibers with cross-linkages covering crystal facies. Reduced from x375.
FIG. 9. Scanning electron micrograph of surface structure of calculus seen in figure 1, A shows microspherule (center). Reduced from x75.
FIG. 10. Scanning electron micrograph of surface structure of calcium phosphate calculus shows abundance of microspherules. Reduced from x 1,500.
FIG. 11. Intracellula r spherules: A , collection of large positive spherules of mucosubstance in renal tubular cells of actively stone-forming patient. PAS stain. B , collection of laminated calcified spherules in renal tubular cells, distinctly visible near lumina of tubules. Alizarin red stain. Intranephronic calculi within lumen of human nephron: C, laminated mucosubstance microlith. PAS, reduced from x 160. D, calcification of same laminated microlith. Alizarin red, reduced from x 155. Randall's plaque and Carr's bodies: E, submucosal collection of Randall's plaques (arrow) near t ip ofrenal papilla. Alizarin red stain. F, scanning electron micrograph of Carr's intralymphatic microlith. Reduced from x 50.
340
341
ULTRASTRUCTURE AND GENESIS OF CALCULI
what this union takes place within the tubular cel.i or in the abnormal pelviocaliceal urine can only be elucidated by further research. Mr. James W. Willard assisted in these studies. Figures 1 to 10 reprinted with permission. 3 Figure 11, Falso reprinted with permission from Carr, R. J.: Aetiology of renal calculi: microradiographic studies. In: Renal Stone Research Symposium. Edited by A. Hodgkinson and B. E. C. Nordin. London: J. & A. Churchill, Ltd., pp. 123-132, 1969.
7. 8. 9.
10.
kinson and B. E. C. Nordin. London: J. & A. Churchill, Ltd., pp. 181-189, 1969. Boyce, W. H. and King, J. S., Jr.: Present concepts concerning the origin of matrix and stones. Ann. N.Y. Acad. Sci., 104: 563, 1963. Keutel, H.J., King, J. S., Jr. and Boyce, W. H.: Further studies of uromucoid in normal and stone urine. Urol. Int., 17: 324, 1964. Malek, R. S. and Boyce, W. H.: Intranephronic calculosis: its significance and relationship to matrix in nephrolithiasis. J. Urol., 109: 551, 1973. Meyer, J. L., Bergert, J. H. and Smith, L. H.: Epitaxial relationships in urolithiasis: the calcium oxalate monohydrate-hydroxyapatite system. Clin. Sci. Mo!. Med., 49: 369, 1975.
REFERENCES
1. Boyce, W. H.: Organic matrix of human urinary concretions. Amer.
J. Med., 45: 673, 1968. 2. Vermeulen, C. W. and Lyon, E. S.: Mechanisms of genesis and growth of calculi. Amer. J. Med., 45: 684, 1968. 3. Boyce, W. H.: Some observations on the ultrastructure of "idiopathic" human renal calculi. In: Urolithiasis: Physical Aspects. Edited by B. Finlayson, L. L. Hench and L. H. Smith. Washington, D.C.: National Academy of Sciences, pp. 97-114, 1972. 4. King, J. S., Jr. and Boyce, W. H.: Amino acid and carbohydrate composition of the mucoprotein matrix in various calculi. Proc. Soc. Exp. Biol. Med., 95: 183, 1957. 5. King, J. S., Jr. and Boyce, W. H.: Analysis of renal calculous matrix compared with some other matrix materials and with uromucoid. Arch. Biochem. Biophys., 82: 455, 1959. 6. Boyce, W. H.: Macromolecular components of kidney calculi in urine. In: Renal Stone Research Symposium. Edited by A. Hodg-
COMMENT This report makes us mindful that stone research has gone a full cycle. In the 1950s Butt advanced the notion of protective colloid in the treatment of urinary stone patients, and Boyce and associates presented their pioneer work on characterizing stone matrix. During the 1960s work on the crystalline aspects of urolithiasis became predominant and work on macromolecules as they relate to urolithiasis was largely neglected except by a few workers. Now with a better understanding of the physical chemistry of crystals and electrolyte solutions there is a resurgence of interest in the relation of urinary macromolecules to urolithiasis. It is very much to be desired that work in this area will now advance pari passu with our increasing understanding of the crystalline features of stones. Birdwell Finlayson University of Florida College of Medicine Gainesville, Florida