J. R. Smid

Mouse Cellular Cementum is Highly Dependent on Growth Hormone Status

Cementum is known to be growth-hormone (GH)-responsive, but to what extent is unclear. This study examines the effects of extremes of GH status on cementogenesis in three lines of genetically modified mice; GH excess (giant), GH antagonist excess (dwarf), and GH receptor-deleted (GHR-KO) (dwarf). Age-matched mandibular molar tissues were processed for light microscope histology. Digital images of sections of first molar teeth were captured for morphometric analysis of lingual root cementum. Cross-sectional area of the cellular cementum was a sensitive guide to GH status, being reduced nearly 10-fold in GHR-KO mice, three-fold in GH antagonist mice, and increased almost two-fold in giant mice (p < 0.001). Cellular cementum length was similarly influenced by GH status, but to a lesser extent. Acellular cementum was generally unaffected. This study reveals cellular cementum to be a highly responsive GH target tissue, which may have therapeutic applications in assisting regeneration of the periodontium.

Mouse Molar Dentin Size/Shape is Dependent on Growth Hormone Status

Growth hormone (GH) status affects dental development, but how GH influences tooth size/shape is unclear. Since GH affects dental epithelial proliferation, we hypothesized that GH influences the tooth crown and root dimensions. Dentin matrix dimensions were measured in longitudinal sections of decalcified first mandibular molars from 3 genetically modified mice: giant (GH-Excess) mice and dwarf (GH-Antagonist and GH-Receptor-Knockout) mice. GH status was found to influence crown width, root length, and dentin thickness. Analysis of these data suggests that GH influences both tooth crown and root development prior to dentinogenesis as well as during appositional growth of dentin. This is concordant with the expression of paracrine GH and GH receptors during tooth bud morphogenesis, and of GH receptors in the enamel organ, dental papilla, and Hertwig’s epithelial root sheath during dentinogenesis. Based on prior studies, these GH morphogenetic actions may be mediated by the induction of both bone morphogenetic protein and insulin-like growth factor-1 expression.

Ameloblast apoptosis and IGF-1 receptor expression in the continuously erupting rat incisor model.

Enamel-producing cells (ameloblasts) pass through several phenotypic and functional stages during enamel formation. In the transition between secretory and maturation stages, about one quarter of the ameloblasts suddenly undergo apoptosis. We have studied this phenomenon using the continuously erupting rat incisor model. A special feature of this model is that all stages of ameloblast differentiation are presented within a single longitudinal section of the developing tooth. This permits investigation of the temporal sequence of gene and growth factor receptor expression during ameloblast differentiation and apoptosis. We describe the light and electron microscopic morphology of ameloblast apoptosis and the pattern of insulin-like growth factor-1 receptor expression by ameloblasts in the continuously erupting rat incisor model. In the developing rat incisor, ameloblast apoptosis is associated with downregulated expression of the insulin-like growth factor-1 receptor. These data are consistent with the hypothesis that ameloblasts are “hard wired” for apoptosis and that insulin-like growth factor-1 receptor expression is required to block the default apoptotic pathway. Possible mechanisms of insulin-like growth factor-1 inhibition of ameloblast apoptosis are presented. The rat incisor model may be useful in studies of physiological apoptosis as it presents apoptosis in a predictable pattern in adult tissues.

A histochemical study of the effects of high doses of sodium fluoride on dipeptidyl peptidase II activity in the rat incisor ameloblast

Female Wistar rats, 3 weeks old, were given sodium fluoride in saline solution (isotonic) by intraperitoneal injection at a dose of either 0, 10 or 20 mg per kg body weight. This treatment was given 9 times over 4.5 days. After fixation by perfusion and demineralization in neutral EDTA, hemi-mandibles were sectioned in a cryostat. Sections were stained for dipeptidyl peptidase II activity, using the specific substrate Lys-Ala-MNA and the coupler Fast Blue B for histochemical localization. Staining indicative of dipeptidyl peptidase II was found in the enamel organ of the incisor, particularly in cells of the stratum intermedium and in both secretory and maturation ameloblasts. This staining was markedly reduced in ameloblasts of rats given either 10 or 20 mg sodium fluoride per kg body weight.

Effects of a Single Intravenous Dose of Sodium Fluoride on Plasma Electrolytes and Metabolites in Rats, Rabbits, and Cockerels

This study assessed the effect of a single intravenous dose of sodium fluoride (20 mglkg body wt.) on plasma ionic fluoride and on some other plasma electrolytes and metabolites in rats, rabbits, and cockerels. At any given time following sodium fluoride administration, the plasma ionic fluoride was highest in rabbits and lowest in cockerels. The rate of removal of fluoride from plasma was slower in rabbits as compared with that in the other two species. Plasma sodium, chloride, total protein, albumin, total globulins, and osmolality were not significantly altered by sodium fluoride in any of these three species. However, plasma phosphate (inorganic), urea, creatinine, and glucose were elevated, and plasma calcium was reduced in the rats and the rabbits, but none was significantly altered in the cockerels. The analyses indicated that species variability does exist in fluoride toxicity.

Growth Hormone and Epidermal Growth Factor in Salivary Glands of Giant and Dwarf Transgenic Mice

Epidermal growth factor (EGF) in rat salivary glands is regulated by testosterone, thyroxin, and growth hormone (GH). Salivary glands of 45-day-old giant and dwarf male and female transgenic mice were examined histologically and by immunohistochemistry (IHC) for EGF. Male giants showed no significant differences from wild-type (WT) parotid and submandibular glands. However, their sublingual glands expressed EGF diffusely and strongly in granular cells within the striated ducts, where they were not found in WT mice. Submandibular gland ducts of female WT were different, having individual granular cells strongly positive for EGF and distributed sporadically along the striated duct walls. Neither female GH-antagonist dwarf mice nor GH-receptor knockout mice had any granular cells expressing EGF in any gland. Obvious presence of granular duct cells in the sublingual glands of giant male mice suggests GH-upregulated granular cell EGF expression. Furthermore, absence of granular duct cells from all glands in female GH-antagonist and GH-receptor knockout transgenic mice suggests that GH is necessary for the differentiation of the granular cell phenotype in female salivary glands.

Physiological variations in plasma testosterone in normal adult male subjects measured by a competitive protein-binding technique.

A method of estimating plasma testosterone utilizing a single paper chromatographic procedure and competitive protein-binding is described. The influence of some conditions of sample collection on plasma testosterone has been investigated. Delay in the separation of plasma from cells and the type of anticoagulant used were both without significant effect. There was an approx. two-fold variability in plasma testosterone concn. in samples withdrawn daily, weekly and monthly from four young, normal adult males.

The effect of intravenous sodium fluoride and synthetic salmon calcitonin on plasma total calcium, inorganic phosphate, and ionic fluoride

SummaryVarious doses of sodium fluoride (NaF), salmon calcitonin (CT) and NaF combined with CT were given intravenously to rats (236±2 g). Blood plasma samples were collected at various times up to 24 hours and measured for total calcium (adjusted for variation in plasma albumin), inorganic phosphate, ionic fluoride, urea, and creatinine. Following injection of NaF alone, significant hypocalcemia and hyperphosphatemia was observed. In contrast, CT injections resulted in hypocalcemia and hypophosphatemia. NaF and CT given in combination generally resulted in the hypocalcemia being equal to or greater than that calculated from the simple addition of their individual effects. Significant increases in both plasma creatinine and plasma urea were observed following treatment with NaF alone or with CT and NaF together, whereas CT alone had negligible effect. These results suggest that NaF does not mimic the effects of CT; rather that NaF and CT interact to modify their individual effects. The influence of NaF probably occurs via an effect on kidney function.

A microspectrophotometric analysis of the effect of fluoride on immature enamel matrix protein of rat molar teeth.

SummaryOver a 24-h period, Wistar rats from 4 litters, 6 to 9 days old, were given five intraperitoneal injections of a solution of 0.9% sodium chloride containing sodium fluoride (3 mg F/kg body weight). Within-litter controls were used. All rats were killed by decapitation 2 h after the final injection and the rat heads, cut sagittally, were processed for protein histochemistry.The intensity of staining of the protein in the enamel matrix of the upper jaw molar tooth buds was quantified using the two-wavelength method of microphotometry. A significant increase in the intensity of staining of fluoride-treated tissues over controls was observed with the histochemical methods specific for arginine (P<0.01), tyrosine (P < 0.05), and cysteine (P<0.05). Other histochemical methods specific for amino acid groups failed to show any significant difference between fluoride and non-fluoride-treated enamel matrix.