Marion D. Francis

Diphosphonates inhibit hydroxyapatite dissolution in vitro and bone resorption in tissue culture and in vivo.

Two diphosphonates containing the P-C-P bond, Cl2C(PO3HNa)2, and H2C(PO3HNa)2 retard the rate of dissolution of apatite crystals in vitro. They inhibit bone resorption induced by parathyroid extract in mouse calvaria in tissue culture and in thyroparathyroidectomized rats in vivo.

Diphosphonates Inhibit Formation of Calcium Phosphate Crystals in vitro and Pathological Calcification in vivo

Two diphosphonates containing the P-C-P bond, CH3C(OH)(PO3HNa)2 and H2C(PO3HNa)2, inhibit the crystallization of calcium phosphate in vitro and prevent aortic calcification of rats given large amounts of vitamin D3. The diphosphonates therefore have effects similar to those described for compounds containing the P-O-P bond but are active when administered orally.

Sites and mechanisms of localization of technetium 99m phosphorus radiopharmaceuticals in acute myocardial infarcts and other tissues

This study was performed to elucidate the localization at the cellular level of technetium-99m phosphorus ((99m)Tc-P) radiopharmaceuticals in acute myocardial infarcts and the mechanisms responsible for (99m)Tc-P uptake in acute myocardial infarcts and other tissues. In 20 dogs with proximal left anterior descending coronary arterial ligation for 1-3 days, elevated calcium levels were measured at all sites of increased (99m)Tc-P uptake (acute myocardial infarcts, necrotic thoracotomy muscle, lactating breast, and normal bone); however, a consistent linear relationship between (99m)Tc-P and calcium levels was not observed. A strong correlation (r = 0.95 and 0.99, n = 2 dogs) was demonstrated between levels of (3)H-diphosphonate and (99m)Tc-P in infarcted myocardium. Autoradiographic studies with (3)H-diphosphonate revealed extensive labeling in the infarct periphery which contained necrotic muscle cells with features of severe calcium overloading, including widespread hypercontraction as well as more selective formation of mitochondrial calcific deposits. Autoradiography also demonstrated labeling of a small population of damaged border zone muscle cells which exhibited prominent accumulation of lipid droplets and focal, early mitochondrial calcification. Cell fractionation studies revealed major localization of both (99m)Tc-P and calcium in the soluble supernate and membrane-debris fractions of infarcted myocardium and less than 2% of total (99m)Tc-P and calcium in the mitochondrial fractions; however, electron microscopic examination showed that mitochondria with calcific deposits were not preserved in the mitochondrial fractions. In vitro studies evaluating the role of serum protein binding on tissue uptake of (99m)Tc-P agents demonstrated that, in spite of significant complexing with serum proteins, serum (99m)Tc-P activity retained the ability to adsorp to calcium hydroxyapatite and amorphous calcium phosphate. In vivo studies showed that concentration of human serum albumin (labeled with iodine-131) in infarcted myocardium reached a maximum of only 3.8 times normal after a circulation time of 96 h, whereas (99m)Tc-P uptake was at least 10 times normal after a circulation time as short as 1 h. It is concluded that: (a) (99m)Tc-P uptake in acutely infarcted myocardium, and possibly other types of soft tissue damage, is limited to necrotic and severely injured cells; (b) concentration of (99m)Tc-P results from selective adsorption of (99m)Tc-P with various forms of tissue calcium stores, including amorphous calcium phosphate, crystalline hydroxyapatite, and calcium complexed with myofibrils and other macromolecules, possibly supplemented by calcium-independent complexing with organic macromolecules; and (c) lack of a linear relationship between (99m)Tc-P and tissue calcium levels mainly results from local differences in composition and physicochemical properties of tissue calcium stores and from local variations in levels of blood flow for delivery of (99m)Tc-P agents.

In vitro andin vivo evaluation of anti-calculus agents

Gem diphosphonates, vicinal polyphosphonates and polyphosphates were found to be effective inhibitors of calculus formation in rats. Sodium citrate and trisodium phosphate were found ineffective. Accumulation of calculus was inhibited by inclusion of the phosphonates in the calculus diet or by topical application of aqueous solutions. The anti-calculus effect of phosphonates was topical rather than systemic and was related to the relative effectiveness of these compounds to inhibit crystal growth of hydroxyapatite. A calculus grading method was used and was reproducible among several graders in assessing calculus production and inhibition. Calculus production in weanling animals fed the control diet occurred at a rapid rate under the conditions described, with maximum scores being reached after about 18 days.RésuméDes diphosphonates de gemme, des polyphosphonates voisins, et des polyphosphates semblent constituer des inhibiteurs actifs de la formation du tartre des rats. Le citrate de sodium et le phosphate trisodique sont inactifs. Les dépôts tartriques sont inhibéspar adjonction des phosphonates à l’alimentation ou par application locale de solutions aqueuses. L’effet antitartre des phosphonates est local plutôt que général et semble lié à l’efficacité relative de ces composés vis à vis de l’inhibition de la croissance cristalline de l’hydroxyleapatite. Un indice de tartre est utilisé et s’avère reproductible pour divers examinateurs pour estimer le dépôt et l’inhibition du tartre. La formation de ce dernier chez des animaux sevrés, soumis à une alimentation témoin, est rapide dans les conditions décrites avec des valeurs maximales atteintes au bout de 18 jours.ZusammenfassungGeminale Diphosphonate, vicinale Polyphosphanate und Polyphosphate erwiesen sich als wirksame Hemmer der Zahnsteinbilding in Ratten. Natriumcitrat und Trinatriumphosphat waren unwirksam. Eine Anhäufung von Zahnstein wurden durch Beigabe der Phosphonate in der Diät oder durch lokale Anwendung von wäßrigen Lösungen verhindert. Die steinhemmende Wirkung von Phosphonaten war eher lokal als systemisch und hing zusammen mit der relativen Wirksamkeit dieser Substanzen bei der Hemmung des Kristallwachstums von Hydroxyapatit. Die Steinmeßmethode wurde verwendent und war bei der Festlegung von Steinproduktion und-hemmung mit mehreren anderen Methoden reproduzierbar. Die Steinbildung bei entwöhnten Tieren, welche die Kontrollnahrung erhielten, erfolgte bei den beschriebenen Bedingungen rasch, wobei die maximalen Werte etwa nach 18 Tagen erreicht wurden.

The development and regression of hypomineralized areas of rat molars

Abstract Hypomineralized areas in enamel which stain with silver nitrate in a manner indistinguishable from incipient carious lesions are present in rats 20–22 days old. The number and severity of these areas is decreased by maintaining the animals on a commercial laboratory chow diet for a period of 70 days, indicating that mineralization of the hypomineralized areas occurs under these conditions. Animals placed on a cariogenic diet showed an increase in number and severity of incipient lesions elaborated by the silver nitrate. The experiment was not designed, however, to distinguish whether some mineralization had occurred in the presence of the active caries formation. Phosphate supplementation of the cariogenic diet resulted in a decrease in number and severity of hypomineralized areas compared with an unsupplemented diet, demonstrating that phosphate either decreases the rate of formation and progress of lesions or favours mineralization. The possibility that topically applied agents such as fluoride may have a mineralizing, as distinguished from a cariostatic, action is discussed.

The effect of phosphanates on dental enamelin vitro and calculus formationin vivo

Di- and poly-phosphonic acids react with polished human dental enamel at acidic and basic pH values to form protective films on the surface which limit the destructive dissolution that can occur in enamel due to acid buffer or calcium chelation effects. The protective effect of these films is demonstrable by electron microscopy. Trisodium ethane-1-hydroxy-1,1-diphosphonate (Na3EHDP) or trisodium methanediphosphonate (Na3MDP) included in a calculus-producing diet significantly reduced the accumulation of calculus on the teeth of rats. The mechanism of calculus prevention appears to be interference with crystal growth of hydroxyapatite as determined by electron microscopy and electron diffraction. From these and other data phosphonates appear to form films on enamel which reduce dissolution of the enamel and also provide a surface which calcifies poorly to limit calculus formation.RésuméDes acides di et polyphosphoniques réagissent avec de l’émail humain poli à des valeurs acides et basiques de pH pour former un film protecteur à la surface qui diminue la dissolution destructrice pouvant se produire dans l’émail par suite des effets tampon acide ou de chélation du calcium. L’effet protecteur de ces films est confirmé par le microscope électronique. Du trisodium éthane-1-hydroxyle-1,1-diphosphonate (Na3EHDP) ou du trisodium méthanedi-phosphonate (Na3MDP), mélangé à un régime produisant du tartre, réduit significatvement l’accumulation de tartre sur les dents de rats. Le mécanisme de prévention du tartre semble devoir interférer avec la croissance cristalline de l’hydroxyleapatite, ainsi que le montrent la microscopie et la diffraction électroniques. D’après ces résultats, les phosphonates semblent former des films sur l’émail qui diminuent la dissolution de l’émail et donnent une surface qui se calcitie mal pour former du tartre.ZusammenfassungDi- und Polyphosphonsäuren reagieren mit poliertem menschlichem Zahnschmelz bei sauren und basischen pH-Werten und bilden Schutzfilme auf der Oberfläche, welche die destruktive Auflösung begrenzen, die im Schmelz durch sauren Puffer oder durch Chelatkomplexbildung mit Calcium erfolgt. Die Schutzwirkung dieser Filme kann durch Elektronen-Mikroskopie nachgewiesen werden. Trinatrium Aethan-1-hydroxy-1,1-Diphosphonat (Na3EHDP) oder, Trinatrium Methandiphosphonat (Na3MDP), welche einer Zahnstein-bildenden Diät zugegeben wurden, verminderten signifikant die Zahnsteinablagerung auf Ratten-zähnen. Der Mechansmus, der für die verminderte Zahnsteinbildung verantwortlich ist, scheint auf einer Interferenz im Kristallwächstum von Hydroxyapatit zur beruhen; dies wurde durch Elektronen-Mikroskopie und Elektronen-Diffraktion nachgewiesen. Diese und andere Befunde zeigen, daß Phosphonate auf dem Schmelz Filme zu bilden scheinen, welche die Auflösung von Schmelz vermindern und zu einer Oberfläche führen, die nur schwach verkalkt und somit die Zahnsteinbildung begrenzt.

The effect of enamel fluoride on acid production by Lactobacillus casei.

Abstract The effect of enamel fluoride on acid production by Lactobacillus casei was estimated using microsystems in which a large area of enamel surface was exposed to a small volume of metabolizing cells, simulating the tooth-plaque relationship. Acid production by L. casei was inhibited by fluoride released from normal and decalcified enamels treated with sodium fluoride or stannous fluoride. Mottled enamel, containing fluorapatite, also served as a source of fluoride to inhibit acid production by L. casei . These data suggest that inhibition of glycolysis by fluoride released from the enamel surface may be responsible, in part, for caries reduction following water fluoridation or the topical application of fluoride.

Inorganic tin: Chemistry, disposition and role in nuclear medicine diagnostic skeletal imaging agents

Abstract A brief review of the chemistry of tin(II) is made with particular emphasis on oxidation and hydrolysis. Spurious oxidation and hydrolysis of tin(II) are reactions encountered in conjunction with the radionuclide technetium which can be important in scintiscan quality and interpretation under use conditions in nuclear medicine. Means to block these reactions are discussed. Some reactions of tin(II) and phosphonates of general interest are examined. The toxicity and biological handling of both tin(II) and tin(IV) are reviewed. Recent methods of reducing and complexing technetium without the use of tin(II) are discussed.

Factors Affecting Uptake and Retention of Technetium-99m-Diphosphonate and 99m-Pertechnetate in Osseous, Connective and Soft Tissues

The bone scanning complex,99mTc−Sn·EHDP, consisting of the nuclide technetium-99m, stannous ion and ethane-1-hydroxy-1,1-diphosphonate, administered intravenously is retained in soft tissues in proportion to increasing calcium content of the tissues. Within bone tissue, the retention is proportional to vascularity and to surface area of calcium phosphate in bones and not necessarily to calcium and phosphate concentration. The nuclidic agent99mTcO4− but not the99mTc-diphosphonate is selectively taken up by the thyroid and this uptake can be blocked by administering sodium perchlorate. Among the connective tissues studied, the tracheal cartilage seems to have the greatest potential to calcify with increasing age of the animal and man. Soft tissue does not retain the bone scanning complex99mTc−Sn·EHDP but does retain99mTcO4−.

Animal Calculus: Methods of Evaluation and of Dietary Production and Control

Twenty-five diets were studied in a search for one that would induce an assessable, reproducible level of calculus in rodents. A diet consisting of 50% cornstarch, 32% non-fat dry milk, 3% liver powder, 5% celluflour, 1% cottonseed oil, 5% powdered sucrose, 1% calcium chloride dihydrate, 2.7% sodium dihydrogen phosphate monohydrate, and 0.3% magnesium sulfate is suggested for future investigations of anti-calculus agents in rats.