Joseph C. Muhler

Effect of gonadectomy and sex hormones on the structure of the rat salivary glands.

R ECENTLY, a number of independent investigators have demonstrated an apparent relationship between the endocrine system and the salivary glands in the experimental animal. Because of the pronounced effect of salivary gland function on dental caries, it is surprising that this relationship has not received more attention. Many of the previously published papers have dealt with the relationship between the sex organs and the salivary glands. In 1940, Lacassagnel pointed out a difference between the histologic structure of the male and female mouse submaxillary glands. It was indicated that the female submaxillary gland assumes the appearance of the male submaxillary gland when the animal receives testosterone and that, further, the male submaxillary gland resembles the female gland in structure when the male animal is treated with folliculin. Lacassagne suggested that these variations observed in the structure of the glands might correspond to differences in the quality of saliva excreted by the two sexes. The same author2 has shown a difference in the histologic structure of the submaxillary gland of pregnant and virgin mice, and has suggested that these glands may be used to test the activity of various hormones, either estrogens or androgens. Lacassagne3 later reported that the submaxillary gland of the rat showed the same reactions as the mouse to the various sex hormones. Lacassagne and Chamorro4 have reported atrophy of the submaxillary gland tubules after hypophysectomy and that repair of these tubules accompanied administration of androgenic hormone. Buillard and Delsuc5 reported that in the parotid and retrolingual glands of the rat there are differences in the size of the cells and the diameter of the acini after gonadectomy and injection of testosterone, which were coincident with those observed by Lacassagne in the submaxillary gland. Working from a different approach, Katagiri and Higashijo6 reported that when the parotid glands are ligated and the submaxillary glands removed in rats there results, among other things, a hypertrophy of the uterus and adrenal cortex, and an atrophy of the testes. Further work by Ginn and Volker7 indicates a pronounced rusting of the fur of female rats, but of males to a much lesser extent, upon removal of the salivary glands. That the salivary glands themselves may have an endocrine function has been suggested by some workers (Ogata8 and Hukusima9 among others), but satisfactory proof is lacking. As yet unexplained is the diminution of weight

ENDOCRINE INFLUENCES UPON THE SALIVARY GLANDS

Evidence is accumulating gradually that the salivary glands, at least in the experimental animal, are associated very intimately with various endocrine organs and that a functional disturbance of one of the latter may induce significant morphologic and physiological alterations in the salivary glands. Much of the early work concerned with the hormonal influences of the salivary glands was that of the Japanese, who published numerous papers speaking particularly of “the internal secretion of the salivary glands” (for example, Ogata, 1934; Takizawa et al., 1938, 1939; Hukusima, 1940). The collective studies of the Japanese investigators are not entirely convincing in affirming that the salivary glands possess an endocrine function in themselves, although some of their work is quite suggestive. Katagiri and Higashijo (1940) reported that ligation of rat parotid gland ducts and extirpation of the submaxillary glands resulted in atrophy of the testes and hypertrophy of the uterus and adrenal cortex. The salivary glands themselves have not been transplanted successfully from their usual location to another site in the body. Ota (1937) has reported that when the salivary glands of dogs were transplanted into bone marrow, the gland elements disappeared, but the ducts of the parotid gland persisted for some time. An extensive report has been published by Ito (1954) on biochemical studies of a “purified salivary gland hormone” called parotin. The French workers have made some of the most significant contributions in attempting to relate the salivary glands and endocrine organs. In the wellknown papers by Lacassagne (1940 a, b, c), i t was shown that there were histological differences between the male and female mouse submaxillary glands and that, if male sex hormone were administered to a female mouse, its submaxillary gland assumed the characteristics of that of the male. Lacassagne suggested that these organs could be used as a test of activity for androgens and estrogens, and speculated that, since a variation in submaxillary gland structure between sexes existed, a corresponding difference in the quality of the saliva excreted also might exist. Subsequently, Raynaud and Rebeyrotte (1949 a, 6 ) did report a difference in the amylase activity of mouse saliva between males and females. Junquiera and his co-workers (1948), however, were unable to find differences in amylase activity of male and female mouse salivary gland tissue extracts, although differences in the protease activity were noted. Buillard and Delsuc (1941) also have reported atrophic changes in the salivary glands following castration. Raynaud (1950) has shown that the injection of testosterone into the submaxillary glands of mice resulted in a marked hypertrophy of the tubules and interpreted this as evidence of a direct action of this adrogen on the gland.

Solubility of Enamel as Affected by Fluorides of Varying pH

Research in the field of enamel solubility is of vital importance because it is known that solubility is a contributing factor in dental caries. There seems to be little doubt today that the use of fluorine compounds are a definite aid in reducing enamel solubility, and thus lowering the incidence of dental caries. The literature contains much evidence in support of this belief, both from a clinical standpoint and from experiments performed in vitro. Miller found that individual teeth appeared to have different rates of decalcification, i.e., tooth solubility, (1) while Pickerell observed that teeth varied in resistance to acids (2). The latter found that the enamel of unerupted teeth was twice as soluble as that of erupted teeth. Dobbs concluded that enamel hardness and resistance to acid decalcification are definitely associated (3). Karshan and Rosebury (4) have studied the relationship of pH, calcium and phosphorus to the solubility of powdered enamel with lactic, succinic, and malic acids. They stated that the pH change at each time interval is definitely proportional to the dissolved phosphorus at that level. McClelland observed that the decalcification occurs very rapidly in hydrogenion concentrations within the ranges of pH 1.0 to 2.5, because at these intensities of acidity a marked loss of weight was obtained within 30 minutes (5). Many other studies dealing with enamel solubility and the influence of various acids, phosphorus and calcium levels, could be cited. It has been shown that, within certain limits, where there is an increased fluorine-content in the water supply, there is a greater resistance to dental decay. Evidence submitted from diverse sources shows that when fluorine is added to the diet or the drinking supply of laboratory animals there is a marked reduction in experimental dental decay. The effectiveness of fluorine as a prophylactic measure in preventing dental caries has been demonstrated both in experimental animals (6), and in man (7, 8). Other work (16) has indicated that when there is a greater resistance to tooth decay, there is an increased amount of fluorine in the tooth substance. Simply knowing that fluorine reduces tooth decay is not necessarily enough, for it is equally as important to find the mechanism by which fluorine acts. Only by a better knowledge of its behavior can we make full use of its poten-

A comparison of the polishing properties of human and bovine enamel.

Although correlations between the abrasion characteristics of human and bovine teeth have been reported (Whitehurst et al., J Oral Ther 4: 181, 1968 and Stookey and Muhler, J Dent Res 47:524, 1968), no studies have compared the polishing properties of human and bovine enamel. Human maxillary incisors have been used in several laboratory studies involving enamel polish, but due to inherent problems associated with their size, curvature, and availability, bovine incisors have been utilized more recently by some investigators (e.g., Barnes et al., J Dent Res 50:966, 1971 and Putt et al., J Dent Res 54:527, 1975) for the in vitro evaluation of polishing agents or pastes. The purpose of this investigation was to compare the polishing properties of human and bovine enamel after treatment with various polishing agents. Extracted human maxillary central incisors and excised bovine permanent incisors were used. Each crown was mounted in a 2 cm-square metal block, and the exposed labial surface was leveled with a mechanical surface grinder as described previously (Putt et al., ibid.). The exposed enamel surface was dulled in 0.2M HCl for 30 sec, thoroughly rinsed, and then brushed for 2,000 strokes at a pressure of 300 gm with a mechanical toothbrushing machine equipped with medium nylon-bristled toothbrushes. Seven dentifrice-grade abrasives were examined as slurries consisting of 25 gm of abrasive and 50 gm of 1% sodium carboxymethyl cellulose solution. The degree of polish was measured before and after treatment by means of a reflectometer system (Putt, Thesis, Indiana University, 1979), whose scores are directly proportional to the level of polish of the tooth specimen. Six replicates were run with each abrasive. The results, which are provided in the Table, -demonstrated that the seven dentifrice-grade abrasives varied widely in their polishing ability, and each abrasive produced similar polish scores with

Experimental Dental Caries III. The Effect of Estradiol and Diethylstilbestrol on Dental Caries, Fluorine Metabolism, and the Salivary Glands of Intact and Gonadectomized Rats

THE studies to be reported here are a portion of the series which has been designed in an attempt to determine how the endocrine organs and their various products affect experimental dental caries and to clarify the relationship of these organs to the salivary glands and fluoride metabolism. Considerable evidence exists1` to warrant further investigation of this problem. Junqueira, Rubinovitch, and Fajer4 have shown enzymatic differences in the salivary glands of male and female mice. Sreebny5 has shown enzymatic alterations as well as histologic changes in the submaxillary gland of the rat following hypophysectomy. Rathje6 has suggested that the action of fluorine in reducing dental caries may be by reduced sympathicotony of the salivary glands mediated through a competition of the thyroid gland for fluorine and iodine essential to its production of hormone, such that a larger quantity of more liquid saliva is produced, thereby increasing the resistance to dental caries. The studies reported from this laboratory have not been designed to measure quantitatively such differences produced by gonadectomy, desalination, and the administration of sex hormones, but rather to determine initially whether alterations of various types occur. Future investigations may be channeled along more exacting lines after these facts have been established, with the aid of conventional or newer and more critical technics.

Salivary Gland Function in the Rat I. Flow, Viscosity, and ph in the Normal and Duct-Ligated Rat

R EPEATED studies from this laboratory have suggested a significant relationship between experimental dental caries in the rat and the function of various endocrine organs, especially the thyroid gland.1-6 The mechanism of this relationship is not known, although convincing evidence suggests that the salivary glands are directly involved. The experimental basis for implicating the salivary glands are changes known to occur in both the histologic structure as well as in the enzymatic activity of these glands concomitant with thyroid dysfunction. These findings have been reported previously.3 6 Undoubtedly, any alteration in histologic structure of an organ must be reflected in an aberration from normal function of that organ. The estimation of dysfunctions in certain organs is relatively simple and may be adapted to experimental investigations in even small laboratory animals. In other organs, especially where a diversity of function exists, the estimation of changes from the normal may be difficult. Because of the almost certain relationship between endocrine dysfunction and salivary gland activity, and the obvious implications regarding dental caries activity, it became of interest to investigate salivary function. The present investigation deals with a series of base-line studies designed in order to determine the accuracy, reproducibility, and applicability of salivary function tests (flow, viscosity, pH) to current endocrine studies in this laboratory.

Sodium Fluoride Reagent Concentration and Powdered Enamel Solubility

NE of the phases of tooth decay, that is, cavitation, for a long time has been assumed to be a dissolution of the tooth mineral material by a weak organic acid, presumably lactic acid. Although direct experimental proof of the assumption is lacking, it is difficult to see at present how it could be otherwise. The effect of fluorine in caries inhibition in human beings in the clinic and albino rats in the laboratory has added to, rather than detracted from, this assumption. This is particularly true since laboratory studies have demonstrated that the Mechanism of the effectiveness of fluorides may be through change in solubility of the tissues involved in the disease.`14 The effect of fluorine upon the solubility of calcium phosphates had been studied previous to the time when fluorine first became recognized as an inhibitor of tooth decay.5 On the basis of work carried out with respect to the effect of fluorine on phosphate and bone solubility, it was even predicted that fluorine might reduce the solubility of dental tissues. Volker6 first demonstrated that fluorine did affect tooth tissue solubility. Buonocore and Bibby7 demonstrated that certain reagents other than sodium fluoride reduced enamel solubility and that the effect varied with different reagents. This point was further emphasized by work done by Muhler and Van Huysen.8 As to the optimum concentration of the fluoride solutions used clinically in the inhibition of tooth decay and in the laboratory studies of enamel solubility, there is no unanimity of opinion. Most clinical investigators use one or two per cent solutions of sodium fluoride and believe that the latter is about as high a concentration as is safe to use with topical methods of application. In most of the in vitro studies of enamel solubility, a concentration of 1/500 reagent solution has been used. In addition to the question of fluoride concentration of reagent solutions, there is also the problem of correct pH of these protecting reagent solutions. It was Bibby9 who first pointed out that acidulated reagent solutions were more effective in reducing enamel solubility than were those at, or around, a neutral pH. In his measurements of the effectiveness of various enamel solubility reducing reagents, he used a pH of 4.0. It is proposed that in this study the effectiveness in reducing enamel solubility of various reagent solution concentrations be examined. Since sodium fluoride is the reagent solution most commonly used to inhibit tooth decay and

Physical Characteristics of a New Cleaning and Polishing Agent for Use in a Prophylaxis Paste

A prophylaxis composition containing a sodium-potassium aluminum silicate abrasive system was evaluated in the laboratory in comparison with several different commercial prophylaxis pastes. The in vitro data indicate that the new composition increases enamel polish and decreases abrasion characteristics without sacrifrcing cleaning efficacy when compared with the commercial products tested.

Enamel Dissolution by Various Food Acidulants in a Sorbitol Candy

The carboxylic acids used as food acidulants were evaluated for their effect on enamel dissolution in vitro. In water, the potential of the food acids to demineralize enamel was directly proportional to their acidity. However, in a sorbitol candy, the amount of enamel dissolution was correlated with the potential of the acids to chelate calcium.

Salivary gland function in rats. II. Effect of thyroid function on salivary flow and viscosity.

Summary 1) We determined the effects of thyroid gland dysfunction on salivary flow and viscosity in the rat. Results indicate that propylthiouracil and radiothyroidectomy reduced salivary flow and increased salivary viscosity. This can be reversed and function restored by administration of thyroxine. Testosterone partially restores salivary flow. 2) These data provide one explanation for the mechanism of alteration in dental caries incidence in the rat following disturbance in thyroid function, although the relationship is probably far more complex than this.