The effect of cell environment on obsteoblastic function was described in this review article. Hypocalcemia, hypocalcified dentin and inhibition of bone formation were demonstrated in rats fed a low calcium diet. These phenomena led us to hypothesize that a low-calcium environment must be one of the important factors to induce the inhibition of bone formation. This proposed mechanism was verified in an in vitro experiment. In the rat neonatal femur cultured in a lowcalcium medium, the head size of the femur, full longitudinal length of the femur and total hexosamine content increased. Alkaline phosphatase activity, chondroitin sulfate synthesis and DNA synthesis activity increased, but proline synthesis activity decreased. In the calvarial bone cells, the total calcium, IP3 contents and PKC activity decreased, but the sensitivity of the cell membrane to a stimulation was enhanced. In mouse osteoblastic MC3T3-E1 cells, c-fos mRNA expression increased after the treatment with fetal bovine serum (FBS) or epidermal growth factor (EGF), and the degree of increase was highest with the 30-min treatment period in both FBS and EGF-treated cells. The expression was significantly higher in the low Ca group as compared to the control group (P<0.01). From these results, it was suggested that tha femur and osteoblasts react to restore the normal cell function against a low-calcium environment, the mediating and intracellular signal transduction system, and a transcriptional activity at the gene level.
Bisphosphonates have a P-C-P bond instead of the P-O-P bond of inorganic pyrophosphate that makes them resistant to enzymatic degradation and gives them a high affinity for hydroxyapatite. They are potent blockers of osteoclastic bone resorption and have been successfully used to treat metabolic bone diseases that involve increased bone resorption. It is possible to synthesize a variety of bisphosphonates by substituting the hydrogen on the carbon atom. The pharmacological characteristics and activity varies greatly from compound to compound, ranging from 1 to 10, 000. Some structure-activity relationships have been found, but no clear-cut one has been established yet. There is a general consensus that the inhibition of bone resorption by bisphosphonates is not caused by the inhibition of dissolution of the hydroxyapatite crystal, but is actually caused through a cellular mechanism that is not completely understood. In the present review article, the possible mode of bisphosphonate action was discussed with special reference to: (1) whether bisphosphonates inhibit the function of mature osteoclasts directly or through osteoblasts and (2) whether bisphosphonates inhibit the proliferation or differentiation of osteoclast progenitors to osteoclasts.
Investigators have demonstrated many types of receptors or acceptors for endogenous substances in salivary glands. These suggest that salivary glands contain receptive systems for many drugs. These receptors can be classified into three types based on the property of saliva secretion: (1) receptors involved in fluid secretion, (2) receptors involved in exocytosis for the protein secretion, (3) receptors involved in both types of secretion. The receptors involved in fluid secretion include the group of α1B, M3, NK-1 receptors coupled with IP3; the group of β2 and VIP receptors coupled with cAMP; and the group of P2Z and P2U receptors coupled with the ATP-gated calcium channel. Whereas α1A, β1, VIP, M3, insulin and H2 receptors mediate exocytosis via the cAMP-proteinkinase A pathway. Moreover, the another pathway of diacylglycerol-proteinkinase C has also been demonstrated to be involved in the exocytosis occurring via NK-1 and M3 receptors. Thus, salivary gland cells have many kinds of drug-receptor system. These receptors are all positive to fluid and/or protein exocytosis. Recently our study on the inhibitory regulation of saliva secretion have suggested the existences of GABA and the GABA-synthetic/metabolic pathway, GABA(A) receptors and benzodiazepine (BDZ) receptors of both central and peripheral types, and furthermore, the coupling of GABA(A) and the central type of BDZ receptors. These receptors are involved in decreasing fluid secretion and amylase release elicited by secretagogues. In the future, mechanisms of the intracellular transduction elicited by BDZ or GABA must be clarified.
This review described the effects of several drugs on the formation and the resorption mechanism of tooth and bone. The enamel formed during high fluoride exposure showed marked hypocalcification, and the organic substances appeared on the incisors in these regions. Such changes following fluoride administration appear to indicate an inhibition of the mineral deposition and/or an inhibition of organic matrix withdrawal. The successive bisphosphonate (HEBP) injections reduced both the progress of the mineral deposition and the formation of the enamel matrix. Colchicine showed a remarkable inhibitory effect on the matrix-forming cells of the dentin and enamel in teeth through its striking inhibitory action on the cytoskeletal system and secretary function. The dentin formation in rabbit and rat incisor dentin was reduced markedly. Colchicine induced reparative dentinogenesis in dental pulp and ectopic calcification in the bone marrow. Both sodium salicylate and aspirin reduced dentin formation through a mechanism that reduces the incorporation of collagen precursors into odontoblasts. Sodium salicylate also inhibited the growth of the upper jaw and tibia in rats. The recruitment of osteoclast-like multinucleated cells was inhibited by sodium salicylate in vitro. A pharmacological approach for studying the formation and resorption mechanism of hard tissue is useful tool for developing hard tissue research.
ODU Plaque-susceptible rats (ODUS/Odu) exhibit markedly heavy plaque formation in the lower incisors and develop both periodontal pockets and gingivitis after being fed a commercially available powder diet. These rats have been established as an inbred strain. We have demonstrated that the ODUS/Odu are a very suitable experimental model for studying periodontitis. We already reported about the allelic distribution, changes of plaque formation and body weight, biochemical nature, toxic activity, vascular permeability factor and bradykinin inactivating factor of the plaque, histological and immunological studies, the pH in the periodontal pocket, amount of saliva, IgA in the saliva, salivary kallikrein, the relationship between sialic acid in the saliva and the serum, leukocyte functions (chemotaxis and superoxide anion) in ODUS/Odu, histamine, mast cell, free radicals, superoxide dismutase activities in gingiva and gingival nerve fibers with substance P or calcitonin gene-related peptide, and effect of diabetes. Streptozotocin-induced diabetic ODUS/Odu may be a useful tool for studying the pathological mechanisms in the development of periodontal tissue breakdown in diabetes. ODUS/Odu should help to further establish the utility of this strain as a model for experimental periodontal disease.
In this study, we introduce a cultivation method for mandibular first molars from mouse embryos. We also investigate the in vitro effects of several drugs on tooth formation. Histological observations demonstrated that the tooth germ dissected from 17-day-old embryonic mice and cultivated for 7 days under the conditions described here showed clear mineralization in the dentin and enamel. Alkaline phosphatase (ALP) activity and calcium content in the tooth germ increased developmentally during 7-day cultivation. The increase of calcium was preceded by that of ALP in the germ. The removal of ALP attached covalently to the external surface of plasma membranes by phosphatidylinositol specific phospholipase C disturbed in vitro mineralization in the tooth germ, suggesting that ALP functioning as an ectoenzyme is involved in the physiological process of tooth formation. To elucidate the effects of calcium regulating hormones and several drugs on tooth mineralization, ALP and calcium content were measured and also the tooth germ was observed histologically. The results obtained from the present study suggest that this in vitro system provides a useful tool for investigating both the direct action of drugs on tooth formation and the mechanisms of drug action.
Progressive periodontal disease is characterized by acute progressive lesions of gingival connective tissues, excessive leukocyte infiltration, and occurrence of a characteristic microflora. A variety of proteolytic enzymes derived from oral bacteria and host cells are found in gingival crevices and thought to play an important role in the onset and development of progressive periodontal disease. The anaerobic bacterium Porphyromonas gingivalis has been implicated in the etiology of the disease. Recently, we have purified a novel arginine-specific cysteine proteinase, termed “argingipain”, from the culture supernatant of the organism. The enzyme was shown to have two important abilities related to the virulence of the organism. One is direct association with periodontal tissue breakdown through its abilities to degrade physiologically important proteins such as human collagens (type I and IV) and to evade inactivation by internal protease inhibitors. The other is associated with disruption of the normal host defense mechanisms through its abilities to degrade immunoglobulins and to inhibit the bactericidal activity of polymorphonuclear leukocytes. The virulence of argingipain was further substantiated by disruption of argingipainencoding genes on the chromosome by use of suicide plasmid systems. On the other hand, we have studied roles of host cell-derived proteinases in the periodontal tissue breakdown. Levels of lysosomal proteinases such as cathepsins B, H, L, G and medullasin were determined in gingival crevicular fluid from periodontitis patients and experimental gingivitis subjects by activity measurement and sensitive immunoassay. The results suggested that all of these enzymes would be involved in the development of both gingivitis and periodontitis.
To study the sites of action of cytokines and anti-angiogenic agents, bioassays have been developed to systematically investigate not only the entire process of angiogenesis in vivo but also its individual steps including capillary tube formation and proliferation of vascular endothelial cells (EC). Angiogenesis in vivo is quantitatively assayed by measuring the carmine dye content in Freund's complete adjuvant-induced mouse pouch granuloma after the intravenous administration of dye solution. Angiogenesis in vitro is quantified by counting the number of microvessels developed from blood vessels cultured in fibrin gel. Tube formation is quantitatively estimated by measuring the total length of capillary tubes from EC cultured in type I collagen gel. EC proliferation is assayed by measuring both the increase in cell number and 3H-thymidine incorporation into the cells. The competence and progression activities in the EC proliferation are analyzed by our convenient method. By these bioassays, the mechanism of an anti-differentiation agent can be easily clarified to develop new types of therapies for rheumatoid arthritis and diabetic retinopathy.