Showing papers in "Calcified Tissue International in 1969"

The inhibition of calcium hydroxypatite crystal growth by polyphosphonates and polyphosphates.

Abstract: The formation of crystalline calcium hydroxyapatite from solutions of calcium and phosphate ions and the inhibition of calcium hydroxyapatite crystal growth by polyphosphonates and polyphosphates have been studied. The polyphosphonates, disodium ethane-1-hydroxy-1,1-diphosphonate and disodium dichloromethane diphosphonate, are effective inhibitors of calcium hydroxyapatite crystal growth. The polyphosphates are also effective inhibitors of calcium hydroxyapatite crystal growth as long as the required level of intact polyphosphate is present in the system. However, because of their hydrolytic instability, which is enhanced by high temperature, low pH, and certain enzymes, the concentration of the polyphosphate decreases with timein vitro, and its activity as an inhibitor is lost. In contrast to the polyphosphates, the polyphosphonates are hydrolytically stable. The polyphosphonates are chemisorbed on the surface of the microcrystallites of calcium hydroxyapatite and, in the manner of other known crystal growth poisons, thus prevent further crystal growth. The stability of the polyphosphonates and their chemisorption on apatite suggest their use in medical and dental applications involving pathological calcium and phosphate metabolism.

The relative contribution of trabecular and cortical bone to the strength of human lumbar vertebrae.

Abstract: An experiment was performed to determine the relative contribution of the cortical shell and of the central trabecular bone to the peak, non-destructive compressive strength of excised human lumbar vertebrae. The vertebral “units” tested utilized the adjacent intervertebral discs to distribute the loads. Among other results the study indicated that (1) the cortex generally contributes 45–75% of the peak strength, regardless of percent ash or physical density of the trabecular bone; (2) when the ash content of a vertebral trabecular bone is 40% of the forces are transmitted via the central trabeculae and, as would be expected, (3) less force is transmitted by way of the central trabeculae in older subjects than in those ≦40 years of age.

An electron microscope study of the formation of the nacreous layer in the shell of certain bivalve molluscs.

Abstract: An electron-microscopic study was made of nacreous shell growth in several species of marine molluscs. Studies of sections of mantle-shell preparations show that the first step in crystal formation is the “polymerization” of part of the pallial fluid to form lamellae parallel to the surface of the epithelium. These lamellae form compartments enclosing a modified apallial fluid. Initiation of crystals occurs in these compartments in contact with a crystal in an adjacent layer. During crystal growth the organic matrix present in the compartment is displaced by the growing surface of the crystal. When growth is complete the crystal is entirely enveloped by a delicate organic sheath. These studies show that the pallial fluid with its organic constituents is responsible for supplying a matrix or substrate for crystal initiation and growth. It serves as a regulatory device for guiding the orderly growth and arrangement of crystals and, further, it may participate in the induction of new crystals. The formation of compartments during shell growth accounts for the uniform thickness, preferred exhibited orientation and mineralogy of the crystals as well as other features exhibited by the mature nacre.

The cancellous bone of the iliac crest.

Abstract: A study of the thickness and spacing of bony structures in the cancellous bone of the iliac crest. Ground sections of methacrylate-embedded material from 67 normal subjects, from 20 to 90 years of age, were examined. Increasing age is accompanied by an increase in the spacing between the bony structures, but not by any significant reduction in their thickness. The thinnest bony structures appear to be undergoing the most remodelling, as judged by the extent of formation and resorption surfaces.

A sensitive in vitro method for studying the induction and inhibition of bone resorption.

Abstract: A sensitive method has been developed for studying the early effects of hormones and other agents on the movement of calcium into and out of bone. Half-calvariae from 6-day-old mice that have been pulsed four days previously with45Ca, are explanted into separate dishes of liquid medium and maintainedin vitro; one half serves as control and the other for experimentation. The time course of action of any agent is followed by removing small samples aseptically from the media and analysing for isotope. Both parathyroid hormone and vitamin A cause extensive bone resorption, and as compared with controls an increased release of45Ca from the treated bones into the medium can be detected within 2 hours. Calcitonin rapidly inhibits the release of45Ca from resorbing bones; the time course is similar to that for its actionin vivo in lowering serum calcium. Our results indicate that agents that induce bone resorption increase both the number of multinucleate osteoclasts and the effectiveness of the existing cells in mobilising calcium. Calcitonin prevents the formation of new multinucleate osteoclasts, and also prevents existing osteoclasts from mobilising bone mineral. Calcitonin inhibits the release of45Ca from living explants maintained in control medium. This reduction is attributed to the suppression of the endogenous resorption that is in progress when the bones are explanted; exchange of isotope still occurs, as in a dead explant.

Mitochondrial Granules in Chondrocytes

Abstract: Rat and mice epiphyseal growth plates were studied with the electron microscope. A gradient of mitochondrial electron-dense granules was found. Chondrocytes in the proliferative zone had few granules, while those of the succeeding zones showed a gradual increase in number and density until the zone of provisional calcification was reached. This zone showed a peripheral distribution of mitochondria and a decrease in the number and density of mitochondrial granules. Isotopic47calcium was used autoradiographically to determine the location of calcium in these cells. Grains were found over the endoplasmic reticulum membranes and over most mitochondria. The demonstration of a gradient of these granules and their spatial relation to the mineralization front suggests a possible involvement of mitochondria in the onset of matrix calcification.

The influence of the periostracum on the shell structure of bivalve molluscs.

Abstract: The periostracum is an outer, quinone-tanned protein layer of the bivalve shell. The conchiolin interprismatic walls of the prismatic outer layer of some bivalves are in structural continuity with the periostracum. It is suggested that calcium carbonate deposition begins so close to the margin that all the periostracal material has not polymerised by the time the first crystals of calcium carbonate are precipitated. Consequently, the periostracal protein is pushed to the edge of growing carbonate crystals, preventing their further lateral growth. Growth proceeds in a vertical direction, as columns, until the influence of the periostracum has waned. Confirmatory evidence is provided by natural-regneration periostracum sheets formed on the inner layers of the shell, which are associated with thin prismatic layers.

Fluoride and bone.

Abstract: Introduction The inorganic compounds of fluorine are widely distributed in nature; high concentrations are found in volcanic rock and, especially, in rock containing phosphates ; the erosion of these rocks is responsible for the presence of fluorides in soil, fresh water and the sea. Fluorine compounds are used in industry, and their use is increasing. Several common drugs, including anaesthetics, contain fluorine, and sodium fluoride has recently been employed in the t reatment of rarefying bone disease. The addition of fluoride to domestic water supplies, as a prophylactic against dental caries, has enhanced the importance of fluoride considerably, and has been the impetus to a world wide investigation of its effects. Despite the large volume of research, however, the effects of fluoride on bone are complicated and are still not fully understood. The first investigation of fluoride toxicity was made by RABUTEAU in 1867 (ROttOLlVl, 1937), and BRANDL and TAPPEINEIr published the results of chronic poisoning in a dog in 1891. The first indication of the importance of fluoride in toxicology came in 1912, when BARTOLUCCI noticed a cattle disease with features resembling osteomalacia in the vicinity of an Italian superphosphate factory and suggested that it might be related to the fluoride in the waste products from the factory. Similar findings were reported by Ct~ISTIA~I (1926), near an alumininm factory in Switzerland, and again by SLAOSVOID in 1934, in the vicinity of a Norwegian aluminium factory. In the same year, RoI~onM identified a disease called \"gaddur\" , which had been known to exist for over a thousand years in volcanic areas of Iceland, as skeletal fiuorosis. Subsequently, thousands of papers have been published, and still are, describing endemic and industrial fluorosis.

A zonal analysis of inorganic and organic constituents of the epiphysis during endochondral calcification

Abstract: A dissection procedure has been devised to permit zonal analysis of the epiphyseal plate of fetal calf leg bones. Samples of whole and washed tissue from the various zones were analyzed for their content of electrolyte and organic constituents, as well as for density, ash and moisture.

Further investigation on the organic-inorganic relationships in calcifying cartilage.

Abstract: The organic/inorganic relationships in calcifying cartilage have been studied in tibialfemoral epiphyses of 5-day-old rats and in costochondral junctions of 1-month-old guinea pigs. The main results are:

Recent progress in the chemistry, crystal chemistry and structure of the apatites.

Abstract: Introduction During the last few years, there has been considerable progress in understanding some aspects of the crystal chemistry and structure of the apatites. As evidence of this, it can be cited that there have been three conferences during the last two years which have been partly, or entirely, devoted to the study of the crystal chemistry and crystallography of the apatites: at Toulouse, France in May 1967; at Princeton University, U.S.A., in May 1968; and at Gaithersburg, U.S.A. in September 1968. This paper is an attempt to review and to consolidate some of this new information as it affects those interested in the structure of the mineral component of bones and teeth. Much of the difficulty which is experienced in investigating the apatites is caused by the inability to prepare single crystals of model compounds for X-ray diffraction studies. I t is therefore appropriate to preface a discussion of the structures of the various model compounds with a consideration of the methods available for the growth of single crystals. If it has not yet been possible to grow single crystals, the discussion is extended to include well-crystallised (and therefore potentially well-eharacterised) samples. At first sight it might seem that many oi the synthetic apatites to be considered are not relevant to the study of hard tissue, but I believe that because the biological apatites are so complicated and have resisted attempts to determine their structures directly, progress will only be made by a wide and detailed study of the chemistry of the apatites in general. The review will begin with two compounds which are distant from the biological apatitcs, namely fluorapatite and chlorapatite. These will be followed by a discussion of the structure of stoichiometric hydroxyapatite, which is the prototype compound of the hard tissues. The complications involved in the nonstoichiometric and carbonate-containing apatites will then be considered, and the review ended with a consideration of the mineral components of bones and teeth.

The effect of some inhibitors on the nucleation and crystal growth of apatite.

Abstract: Mg, Sr and F ions acted as inhibitors of apatite formation when added to a calcification buffer separately or in combinations.

The effect of excessive acid feeding on bone

Abstract: Excessive administration of ammonium chloride to normal adult male rats receiving a diet adequate in vitamin D caused the development of osteoporosis. The osteoporosis was due to loss of bone substance and bone mineral associated with increased bone resorption.

Calcification in sea urchins. I. A tetracycline investigation of growth of the mature test in Strongylocentrotus intermedius.

Abstract: Growth of the test ofStrongylocentrotus intermedius was examined with the aid of tetracycline. In summer specimens, tetracycline fluorescence was not observable in the plates, except in the apical system and its adjacent area. An array of fibrous elements with a milky white autofluorescence was oriented perpendicularly to each suture line. In winter specimens the elements were evident only in demineralized sections. The growth gradient of each plate estimated on winter specimens was as follows: longitudinal sture » inner surface = mammelon and part of boss in tubercle > latitudinal suture > outer surface of base of spine shaft > outer surface of plate itself. The mature test as a whole increases in size by two modes of growth, an addition of new plates to the apical system and an accretion on fixed sites of each plate. The results suggest that fibrous elements are involved in the test growth, and that there is a different mechanism of the test growth between summer and winter specimens. The apical system may be equipped with a specific mechanism of calcification which permits an active formation of new plates throughout the year.

Ionic interaction with bone mineral

Abstract: The effect of Mg2+, citrate3−, F− and SO42− on the exchange of45Ca2+ and32PO43− with the mineral phase of bone (ethylenediamine-extracted) was determined under physiological conditions. Mg2+ substituted for Ca2+ at the adherent liquid film (hydration shell), while citrate3− displaced PO43− from both the adherent liquid film and the crystal surface. These bone-seeking elements thus increased the apparent solubility of bone mineral. In contrast, F− markedly reduced the concentration of Ca2+ and PO43− in both the ambient solution and the adherent liquid film, probably by inducing the formation of a CaF2 boundary layer at the crystal surface. There was a net uptake of Ca2+ and PO43− by the solids phase in the presence of Mg2+ and F−, but not in citrate3− solutions. SO42− did not significantly affect the solubility, dissolution, or growth of bone mineral.

Osteogenesis imperfecta: effect of magnesium administration on pyrophosphate metabolism.

Abstract: The bone collagen of human osteogenesis imperfecta is a potent inhibitor ofin vitro calcification. The inhibition was removed by treatment of the collagen with pyrophosphate in the presence of magnesium ions. Elevated levels of serum and urinary inorganic pyrophosphate were found in 28 patients with osteogenesis imperfecta. Oral administration of magnesium oxide or magnesium sulfate to four of these patients significantly reduced their serum and urinary levels of pyrophosphate.

Enzymes of protein and phosphate catabolism in rat bone. I. Enzyme properties in normal rats.

Abstract: A study was made of several enzymatic activities in rat femur and tibia. Quantitative assays were developed for the determination of acid and alkaline phosphatase, lactate dehydrogenase, proteolytic activity, leucine aminopeptidase, glycylglycine dipeptidase, glutamate dehydrogenase, aspartate transaminase, and alanine transaminase. Some properties of these enzymes were studied using metaphyseal homogenates. The distribution of activity in the metaphysis, diaphysis, and marrow was determined. The activity per mg of DNA was highest in the metaphysis and lowest in marrow. The activities in general were 40–70% lower in the diaphysis than in the metaphysis. Metaphyseal enzymatic activity was the same in 29- and 49-day-old rats but lower in 82-day-old rats. Proteolytic activity at pH 8.0 differed from the other activities studied being higher in marrow than in bone and increasing markedly with age.

Intestinal absorption of calcium-47 and strontium-85 in lactating rats.

Abstract: The absorption of calcium and strontium from the gastrointestinal tract was determined in lactating rats between the 14th and 16th day of lactation using radioactive isotopes of calcium (45Ca and47Ca) and strontium (85Sr) both in thein vivo and in thein vitro experiments.

The mechanism of bone resorption in laying hens

Abstract: The medullary bone in the femora of laying hens and of oestrogen-treated cocks has been examined by histological, histochemical, alpharadiographic and microroentgenographic techniques while undergoing both natural resorption and resorption induced by injection of parathyroid extract. In the hens, natural resorption was brought about by egg-shell formation and in the cocks by withdrawal of oestrogen. Natural resorption was accompained by increasing basophilia, azurophilia and metachromasia and by decreasing alpharadiographic and microradiographic density of the trabeculae of the medullary bone. These changes were observed initially in the central region (towards the marrow cavity) and subsequently in the peripheral regions also, where the number of osteocytes per unit area of bone was less. Parathyroid extract enhanced all these effects. It appears that resorption of medullary bone was brought about by osteocytic osteolysis and by osteoclasis, but it was not possible to determine whether both processes occurred concurrently or whether the osteoclasts attacked the trabeculae only after they had been partially degraded by osteolysis. These results are consistent with the hypothesis that the natural resorption of medullary bone during egg-shell formation in hens and following the withdrawal of oestrogen in cocks is induced by parathyroid hormone.

The effect of polyvalent metal ions on the stability of a buffer system for calcification in vitro.

Abstract: The effect was studied of various polyvalent metal ions on the metastability of a calcification buffer. At concentration of 1 μM or less, Pb2+, Fe2+, Fe3+, Cr3+, Bi3+ and Al3+ ions were found to destabilize the buffer. The results suggest that insoluble phosphates formed by these ions initiate the precipitation of calcium apatite. The implications of these findings to pathological calcification and calciphylaxis are discussed.

The relationship of dietary calcium intake to radiographic bone density in normal and osteoporotic persons.

Abstract: Lifetime daily calcium intake was estimated through interview of 398 individuals from 15 to 90 years of age. The correlation of calcium intake with vertebral mineralization as determined by quantitative radiographic densitometry was low but persistently significant.

Medullary bone of laying hens during calcium depletion and repletion

Abstract: Bone cell activity and the composition of the femur of laying hens were studied during 7 days of calcium depletion on a 0.13% calcium diet and 7 days of calcium repletion on a 3.2% calcium diet. Histologically, only cortical bone showed clear signs of bone resorption and osteoclastic activity during the depletion period. The number of osteoclasts in medullary bone varied little from control values throughout both calcium depletion and repletion, except for a significant increase on the first day of depletion. The major histologicalchange in medullary bone was a marked increase in the number of osteoblasts on the third, fifth and, to a lesser extent, seventh, day of depletion. The number of osteoblasts in medullary bone was positively correlated with its osteoid content and negatively correlated with its degree of calcification. Alkaline phosphatase activity of medullary bone increased with the time the hens had been on the calcium-deficient diet. Returning the hens to the 3.2% calcium ration caused, within one day, a significant increase in medullary bone calcification, a decrease of osteoblast numbers to, or below, control levels, and a drastic reduction in alkaline phosphatase activity of medullary bone. The significance of these findings in relation to the control of bone cell populations and the functions of medullary bone is discussed.

Are mitochondria directly involved in biological mineralisation? The mitochondrion and the origin of bone.

Abstract: The importance of the mitoehondrion in biological calcification as proposed by S~A~mo and G~EENSPA~ (1968) raises the question of the evolutionary reason for the affinity of the mitochondrion for calcium ions. Recent workers now seem agreed tha t the first vertebrates were marine (SrJELDNAES, 1967 ; Dv, NISO~, 1967 ; ItALSTEAD, 1969). Although bone acts as an important calcium store in terrestrial vertebrates, such a role would have been superfluous in the sea, where there is (and was) an abundance of calcium ions. Although URmT (1964) has suggested tha t bone may have evolved as a calcium, as well as phosphate, store, the main problem facing marine animals would have been to get rid of the excess calcium. WESTOLL (1942) proposed tha t the early vertebrates \"unable to cope with all their intake of calcium.. , deposited it wherever there were immobile or inert tissues\". The ability of the mitochondria to accumulate calcium and phosphate ions in the cMcinm/phosphate molar ratio 1.7 (that of apati te being 1.65) and subsequently liberate them, seems to be quite simply an excretionary process to deal with excess calcium ions entering the organism from the environment. This is perhaps a type of \"sodium p u m p \" mechanism enabling the organism to maintain a reasonable degree of homeostasis. There remains the problem of how the ions or mineral are transferred from the mitochondria to the site of mineralisation. In general terms it might be expected tha t the skin would be the ideal site, since this is the main barrier between the internal and external environment of an organism the first line of defence, as it were. Following SHArrao and GRV, V, NSrAN'S suggestion, it now becomes possible to outline a general theory of the origin and early evolution of calcified tissues in the vertebrates. The acquisition of a siliceous skeleton in diatoms, radiolaria and sponges is presumably a quite different matter . The origin of vertebrate calcified tissues was primarily related to p r o b l e m s of osmosis, as HOMER SMITK surmised (1953) although not, as he believed, a waterproofing agent. In all likelihood, such tissues first evolved as a byproduct of calcium excretion (W]~sToLL, 1942). The origin of bone has been linked to the seasonal phosphate cycle in the sea (I-~ALSTEAD TA/~LO, 1964; ItALSTEAD, 1969). I t seems reasonable to suggest that, during the seasonal abundance of assimilable phosphates, calcium could have been readily deposited extracellularly in the form

An electron microscope study of the formation of the ligament of Mytilus edulis and Pinctada radiata

Abstract: An electron microscope study was made of the formation and structure of the inner ligament ofMytilus edulis andPinctada radiata. This part of the ligament is derived from the isthmus cells which are irregular columnar in shape. They exhibit a prominent rough endoplasmic reticulum and a Golgi apparatus, which are concerned with the elaboration of vesicles and granules eventually incorporated into an integral part of the conchiolin.

Calcareous sponge spicules: a study of magnesian calcites.

Abstract: The results of chemical, X-ray diffraction and infra-red analyses, together with specific gravity determinations, are presented for the spicules of seven species of Calcarea. The spicules are composed of magnesian calcite. Traces of quartz were detected, but are ascribed to contamination. Aragonite was sought but not proven, although the presence of quartz complicates its detection. The Mg content varied from 1.25–3.15% between species, and increased with spicule size in the two species studied; it also correlated closely with the displacement of the calcite 112 diffraction peak. Minor constituents estimated were Sr2+, Na+ and SO 4 2− :Sr2+ was relatively constant, the other two variable. Traces of Al3+, Si, Mn2+ and Li+ were also detected. Average specific gravities ranged from 2.58–2.62, smaller spicules having a wider range Composition is considered in relation to spicule secretion and classification of the Calcarea.

A structural basis for the transphosphorylation of nucleotides with hydroxyapatite.

Abstract: The basis for transphosphorylation between nucleotides and hydroxyapatite (HA) has been explored. Using a recently-proposed model for the surface of HA and the structure of 2- and 3-membered polyphosphates, a reasonable atomic explanation can be shown for this reaction. Transphosphorylation has been found to result in a pyrophosphate on HA which is distinctive from pyrophosphate absorbed onto HA from solution. The models suggest that this distinction is due to a different orientation of the pyrophosphate on the surface of the HA depending on the origin of the pyrophosphate.

Electron microscopical and microprobe observations on the cell sheath of stimulated osteocytes.

Abstract: Rats were treated with 0.6 mg (0.1 mg twice a week) of dihydrotachysterol and the following electron microscopic and electron-probe X-ray microanalytical findings were made: 1. The periosteocytic mucopolysaccharide sheath is definitely enlarged in the treated animals. In it were found, in variable quantities and distribution, collections of round or needle-shaped electron-dense particles. 2. By electron-probe X-ray microanalysis, high concentrations of calcium and phosphorus were detected in the cell sheath, where the Ca/P ratio may reach the value of pure apatite.