Calcif. Tissue Int. 29, 181-182 (1979)
Calcified Tissue International ~, 1979 by Springer-Verlag
Presence o f " Crystal Ghosts" in Bone Nodules E. B o n u c c i 1st Institute of Pathological Anatomy. University of Rome. Viale Regina Margherita 324.00161 Rome. Italy M a n y i n t e r e s t i n g c o n c l u s i o n s a r e d r a w n in the pap e r b y M a r t i n o , Y e a g e r , a n d T a y l o r [1], s o m e o t h ers, h o w e v e r , a r e u n a c c e p t a b l e . T h e a u t h o r s w r i t e that in d e c a l c i f i e d b o n e t h e y f o u n d no " ' c r y s t a l g h o s t s , " i.e., o r g a n i c f i l a m e n t s w h i c h h a v e the s a m e s h a p e , size, a n d a r r a n g e m e n t as t h o s e o f ino r g a n i c c r y s t a l s , as d e s c r i b e d b y B o n u c c i et al. [29] a n d as c o n f i r m e d for c a l c i f y i n g t i s s u e s b y m a n y o t h e r s [10-14]. M a r t i n o , Y e a g e r , a n d T a y l o r a l s o
state t h a t the t e r m " ' c r y s t a l g h o s t s " is m i s l e a d i n g a n d s h o u l d be a v o i d e d . T h e y h a v e , u n f o r t u n a t e l y , failed to r e c a l l that c r y s t a l g h o s t s are n e v e r r e c o g n i z a b l e u n d e r the e l e c t r o n m i c r o s c o p e in s p e c i m e n s d e c a l c i f i e d b e f o r e e m b e d d i n g [2, 3, 6-8], a n d t h a t , conversely, they are visible when decalcification o c c u r s a f t e r e m b e d d i n g (Fig. 1). This is p r o b a b l y d u e to t h e i r s o l u b i l i z a t i o n d u r i n g d e h y d r a t i o n a n d s p e c i m e n p r e p a r a t i o n w h e n t h e y are not " ' p r o -
Fig. 1. Left: calcification nodules in cartilage decalcified before embedding; they are surrounded by a lamina limitans and contain amorphous, granular, and filamentous material. Right: calcification nodules in cartilage decalcified after embedding: crystal ghosts are visible in the place of lamina limitans and in matrix of the nodules. Uranyl acetate and lead citrate, x 50,000 and (right) x 80,000 S e n d qff'print reqttests to Prof. E. Bonucci at the above address.
0171-967X/79/0029-018 i $01.00
182
tected" by inorganic substance. The material studied b y M a r t i n o , Y e a g e r , a n d T a y l o r w a s d e c a l c i f i e d f o r at l e a s t 10 d a y s b e f o r e e m b e d d i n g (as w a s t h a t s t u d i e d in t h e w o r k o f T h y b e r g q u o t e d b y t h e m ) , s o t h a t it w o u l d h a v e b e e n a m a z i n g if t h e y h a d s p o t t e d any crystal ghosts. If they had used the correct technique, of course, the crystal ghosts could have b e e n r e l i e d o n to s h o w up. T h e m a i n a i m o f this c o m m e n t is t h e r e f o r e t o point out that. whatever the function and biological significance of "crystal ghosts" may be, the term i t s e l f is n o t m i s l e a d i n g o r d i s r e p u t a b l e , a n d it will b e c o m e s o if it c a n be s h o w n t h a t t h e s e s t r u c t u r e s a r e t e c h n i c a l a r t i f a c t s o r if a b e t t e r n a m e c a n b e found.
References 1. Martino, L.J., Yeager V.L., Taylor J.J.: An ultrastructural study of the role of calcification nodules in the mineralization of woven bone, Calcif. Tissue Int. 27:57-64, 1979 2. Bonucci, E.: Fine structure of early cartilage calcification, J. Ultrastruct. Res. 20:33-50, 1967 3. Bonucci, E.: Further investigation on the organic/inorganic relationships in calcifying cartilage, Calcif. Tissue Res. 3:3854, 1969 4. Bonucci, E.: The locus of initial calcification in cartilage and bone, Clin. Orthop. 78:108-139, 1971 5. Bonucci, E.: The organic-inorganic relationships in bone
E. Bonucci: "'Crystal Ghosts" in Bone Nodules
6.
7.
8.
9.
10.
11.
12.
13.
14.
matrix undergoing osteoclastic resorption, Calcif. Tissue Res. 16:13-36, 1974 Bonucci, E.: The organic-inorganic relationships in calcified organic matrices. In: Physico-chimie et cristallographie des apatites d'int6r~t biologique, pp. 231-245. Colloque n. 230 du CNRS. Paris. 1975 Bonucci, E., Derenzini, M.. Marinozzi, V.: The organic-inorganic relationship in calcified mitochondria, J. Cell Biol. 59:185-211, 1973 Bonucci, E., Gherardi, G.: Histochemical and electron microscope investigations on medullary bone, Cell Tissue Res. 163:81-97, 1975 Bonucci, E., Reurink, J.: The fine structure of decalcified cartilage and bone: a comparison between decalcification procedures performed before and after embedding, Calcif. Tissue Res. 25:179-190, 1978 Appleton, J.: Ultrastructural observations on early cartilage calcification. The use of chromium sulphate in decalcification, Calcif. Tissue Res. 5:270-276, 1970 Appleton. J.: Ultrastructural observation on the inorganic/ organic relationships in early cartilage calcification, Calcif. Tissue Res. 7:307-317, 1971 Blakey. P.R.. Guy, R., Happey, F., Lockwood, P.: The effect of chemical modifications on the morphological structure of keratin, J. Appl. Polymer Sci.. Appl. Polymer Symp. 18:193-199. 1971 Kuhar. K.J., Eisenmann. D.R.: Fluoride-induced mineralization within vacuoles in maturative ameloblasts of the rat, Anat. Rec. 191:91-102. 1978 Smith, J.W.: The disposition of proteinpolysaccharide in the epiphyseal plate cartilage of the young rabbit. J. Cell Sci. 6:843-864, 1970
Calcified Tissue International ~, 1979 by Springer-Verlag
Presence o f " Crystal Ghosts" in Bone Nodules E. B o n u c c i 1st Institute of Pathological Anatomy. University of Rome. Viale Regina Margherita 324.00161 Rome. Italy M a n y i n t e r e s t i n g c o n c l u s i o n s a r e d r a w n in the pap e r b y M a r t i n o , Y e a g e r , a n d T a y l o r [1], s o m e o t h ers, h o w e v e r , a r e u n a c c e p t a b l e . T h e a u t h o r s w r i t e that in d e c a l c i f i e d b o n e t h e y f o u n d no " ' c r y s t a l g h o s t s , " i.e., o r g a n i c f i l a m e n t s w h i c h h a v e the s a m e s h a p e , size, a n d a r r a n g e m e n t as t h o s e o f ino r g a n i c c r y s t a l s , as d e s c r i b e d b y B o n u c c i et al. [29] a n d as c o n f i r m e d for c a l c i f y i n g t i s s u e s b y m a n y o t h e r s [10-14]. M a r t i n o , Y e a g e r , a n d T a y l o r a l s o
state t h a t the t e r m " ' c r y s t a l g h o s t s " is m i s l e a d i n g a n d s h o u l d be a v o i d e d . T h e y h a v e , u n f o r t u n a t e l y , failed to r e c a l l that c r y s t a l g h o s t s are n e v e r r e c o g n i z a b l e u n d e r the e l e c t r o n m i c r o s c o p e in s p e c i m e n s d e c a l c i f i e d b e f o r e e m b e d d i n g [2, 3, 6-8], a n d t h a t , conversely, they are visible when decalcification o c c u r s a f t e r e m b e d d i n g (Fig. 1). This is p r o b a b l y d u e to t h e i r s o l u b i l i z a t i o n d u r i n g d e h y d r a t i o n a n d s p e c i m e n p r e p a r a t i o n w h e n t h e y are not " ' p r o -
Fig. 1. Left: calcification nodules in cartilage decalcified before embedding; they are surrounded by a lamina limitans and contain amorphous, granular, and filamentous material. Right: calcification nodules in cartilage decalcified after embedding: crystal ghosts are visible in the place of lamina limitans and in matrix of the nodules. Uranyl acetate and lead citrate, x 50,000 and (right) x 80,000 S e n d qff'print reqttests to Prof. E. Bonucci at the above address.
0171-967X/79/0029-018 i $01.00
182
tected" by inorganic substance. The material studied b y M a r t i n o , Y e a g e r , a n d T a y l o r w a s d e c a l c i f i e d f o r at l e a s t 10 d a y s b e f o r e e m b e d d i n g (as w a s t h a t s t u d i e d in t h e w o r k o f T h y b e r g q u o t e d b y t h e m ) , s o t h a t it w o u l d h a v e b e e n a m a z i n g if t h e y h a d s p o t t e d any crystal ghosts. If they had used the correct technique, of course, the crystal ghosts could have b e e n r e l i e d o n to s h o w up. T h e m a i n a i m o f this c o m m e n t is t h e r e f o r e t o point out that. whatever the function and biological significance of "crystal ghosts" may be, the term i t s e l f is n o t m i s l e a d i n g o r d i s r e p u t a b l e , a n d it will b e c o m e s o if it c a n be s h o w n t h a t t h e s e s t r u c t u r e s a r e t e c h n i c a l a r t i f a c t s o r if a b e t t e r n a m e c a n b e found.
References 1. Martino, L.J., Yeager V.L., Taylor J.J.: An ultrastructural study of the role of calcification nodules in the mineralization of woven bone, Calcif. Tissue Int. 27:57-64, 1979 2. Bonucci, E.: Fine structure of early cartilage calcification, J. Ultrastruct. Res. 20:33-50, 1967 3. Bonucci, E.: Further investigation on the organic/inorganic relationships in calcifying cartilage, Calcif. Tissue Res. 3:3854, 1969 4. Bonucci, E.: The locus of initial calcification in cartilage and bone, Clin. Orthop. 78:108-139, 1971 5. Bonucci, E.: The organic-inorganic relationships in bone
E. Bonucci: "'Crystal Ghosts" in Bone Nodules
6.
7.
8.
9.
10.
11.
12.
13.
14.
matrix undergoing osteoclastic resorption, Calcif. Tissue Res. 16:13-36, 1974 Bonucci, E.: The organic-inorganic relationships in calcified organic matrices. In: Physico-chimie et cristallographie des apatites d'int6r~t biologique, pp. 231-245. Colloque n. 230 du CNRS. Paris. 1975 Bonucci, E., Derenzini, M.. Marinozzi, V.: The organic-inorganic relationship in calcified mitochondria, J. Cell Biol. 59:185-211, 1973 Bonucci, E., Gherardi, G.: Histochemical and electron microscope investigations on medullary bone, Cell Tissue Res. 163:81-97, 1975 Bonucci, E., Reurink, J.: The fine structure of decalcified cartilage and bone: a comparison between decalcification procedures performed before and after embedding, Calcif. Tissue Res. 25:179-190, 1978 Appleton, J.: Ultrastructural observations on early cartilage calcification. The use of chromium sulphate in decalcification, Calcif. Tissue Res. 5:270-276, 1970 Appleton. J.: Ultrastructural observation on the inorganic/ organic relationships in early cartilage calcification, Calcif. Tissue Res. 7:307-317, 1971 Blakey. P.R.. Guy, R., Happey, F., Lockwood, P.: The effect of chemical modifications on the morphological structure of keratin, J. Appl. Polymer Sci.. Appl. Polymer Symp. 18:193-199. 1971 Kuhar. K.J., Eisenmann. D.R.: Fluoride-induced mineralization within vacuoles in maturative ameloblasts of the rat, Anat. Rec. 191:91-102. 1978 Smith, J.W.: The disposition of proteinpolysaccharide in the epiphyseal plate cartilage of the young rabbit. J. Cell Sci. 6:843-864, 1970
