This is a summery and review aimed at proposing a more consistent interpretation of the matrix vesicle calcification including the current views of the process of overall biological calcification. In this review, I focused on the ultrastructural characteristics of matrix vesicles in various calcifying tissues, and special attention was paid to the recent electron microscopic studies of anhydrously prepared tissues which confirm the assertion that matrix vesicles are the initial sites of biological calcification in various calcified tissues. Then I introduced the pertinent data ralated to the cytochemistry and biochemistry, especially data related to the phosphatases, phopholipids, protein-polysaccharides and another proteins. I synthesized a scheme describing how biological calcification be regulated by the cells during calcified tissue formation mediated by matrix vesicles. That is, skeletal tissue cells control calcification, not only regulating and formation of the organic matrix but also by secreting matrix vesicles at the exact site where calcification will be initiated.
The major theories of matrix vesicle biogenesis, budding from plasma membrane, cell degeneration, exocytotic secretion and subunit self-assembly theory, were also introduced.
Finally, I discussed on the subsequential calcification, that is growth hydroxyapatite crystals being a continuation by direct extention, and secondary nucleation from previously formed seam of calcified tissues.

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A 0.5 per cent acetone solution of 9, 10-dimethyl-1, 2-benzanthracene (DMBA) was treated to the buccal mucons membrane in a hamster three times a week. The results obtained were as follows
1. After 19 weeks of the experimental term, 37 hamsters amoung 37 which were pathologically studied were observed to develop symptoms of carcinomas.
2. Process of experimental production of carcinomas; The buccal mucous membrane treated with DMBA was seen to be ulcerated in 3 to 6 weeks. After 10 weeks, a nodular or papillary tumor was seen to be produced. Such tumors gradually increased in size and carcinomas were noticed to be produced in 19 weeks. These carcinomas were diagnosed as squamous cell carcinoma in the pathological studies.
In conclusion, although, previous reports of the production of carcinomas in the buccal mucosa of hamsters were unsuccessful, we produced the carcinomas in a short time.

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The effects of fosfomycin (FOM) on growth, morphology and the fine structure of Fusobacterium nucleatum cells were examined. The minimal inhibitory concentration (MIC) of FOM against F. nucleatum cells growing exponentially was 1.56μglml. Phase-contrast and scanning electron microscopy showed that the cells were elongated by exposure to 1/2 MIC of FOM, and vesicles were formed on the cells above the MIC. Transmission electron microscopy revealed that the vesicle was consisted of outer membrane only or outer membrane, cytoplasmic membrane and cytoplasm. Other changes in the cell structure than those mentioned above included invagination of outer membrane, cytoplasmic division and protoplast formation.
It seems that the vesicle formation and the invagination of outer membrane are the structural changes related to cell lysis by FOM.

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1) Comparison of demineralization patterns by different pH of acidic gelatin (Fig. 1): The buccal and lingual enamel surfaces of 291 young premolars were painted with varnish leaving a ‘window’. They were placed in acidic gelatin (Silverstone) at pH 3.5, 4.5, and 5.5, for 11-415 days. The piano-parallel ground sections were microradiographed. pH 3.5 produces very intense and deep demineralization lesions with thin remineralized surface layer (Figs. 2 and 3). pH 4.5 and 5.5 produce lesions which can be divided into 5 layers from surface to bottom, (a) highly remineralized narrow surface layer, (b) demineralized layer, (c) highly remineralized narrow layer, (d) narrow demineralized layer, (e) narrow hypermineralized layer (Figs. 4-9). Radiolucency and depth of lesions are pH 3.5>4.5>5.5.
2) Effect of changing pH during demineralization (Fig. 1): pH 3.5→4.5, pH 3.5→5.5, pH 4.5→5.5 The upper layer of lesions shows characteristic lower pH type demineralization, however, lower layer shows higher pH type changes (b-e layers)(Figs. 11-15). pH 4.5→3.5, pH 5.5→3.5 pH 4.5 or 5.5 type changes (a-d layers) are observed at the upper level of lesions and pH 3.5 type intense demineralization is seen at the lower level (Figs. 16-18). pH 3.5→4.5→3.5 At the bottom of deep pH 3.5 type lesion, narrow pH 4.5 type changes (b-d layers) with narrow intensely demineralized deeper layer can be observed (Figs. 19 and 20). pH 4.5→3.5→4.5 pH 3.5 type lesion is sandwiched the bylayers showing pH 4.5 type changes (Fig. 21).
The results of present study indicate that lesions produced by preceding demineralization by higher pH shows very high acid resistance for subsequent demineralization by lower pH and that the influence of changing pH appears mainly at the bottom of lesions rather than at the upper level.

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The replacement of the first premolars in the mandible and maxilla of Lesser Japanese shrew-moles, Dymecodon pilirostris, and in the maxilla of Greater Japanese shrew-moles, Urotrichus talpoides was observed by the examination of skeletal material and X-ray examination of juvenile in-dividuals. On both species of shrew-moles, the second deciduous premolars are notreplaced. Among extant metatherian and eutherian mammals, only tapirs (Tapirus) show the first premolar replacement. A mong extinct mammals, the evidence regarding the replacement of the first premolar has not been reported as yet, except for the Tapiroidea of the Pleistocene (Simpson, 1945) and the Eocene (Radinsky, 1963). Based on the generally accepted concept that the first premolar if present is not replaced in mammals, Ziegler (1971) erroneously identified the non-replaced second premolar of the Japanese shre wmoles as the “first” premolar. Accordingly, only two premolars are present between the “first” pre-molar and the molar series so that the “second” premolar is regarded to have been lost. Anterior to the “first” premolar on the maxilla are four teeth of which first three have been identified as inci-sors and last tooth as canine by Ziegler. Based on the re-examination of skeletal material, it is now clear that only two incisors are on the premaxillary bone in these shrew-moles, and that Ziegler id entified the first tooth of the tooth series on the maxilla as the third incisor. The identification given by Ziegler is antagonistic to the definition of incisors and canines.

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Five dimensions of head and face by Martin's somatometric method, and nine dimensions of dental arch and palate were measured from Formosan aborigines, 95 males aged 13-15 years.
Simple and multiple correlation coefficients among these dimensions were examined. The results may be summarized as follows:
1) There are high positive correlations among breadths of arch, and among lengths of arch. Between breadths and lengths of arch, breadths of anterior arch have heigher positive correlations with lengths of arch than breadths of posterior arch have. Palatal vault height have no correlation with any breadths and lengths of arch.
2) There are significant positive correlations among dimensions of head and face, except for correlation between head length and head breadth. Bizygomatic breadth have expressly high correlations with other dimensions of head and face.
3) Dimensions of head and face have significant positive correlations with breadths of arch and palatal vault height, but not with lengths of arch.
Breadths of arch have expressly high correlations with bizygomatic breadth and bigonial breadth, then they have high correlations with head breadth and morphologic facial height, but not with head l ength.
Palatal vault height has signficant correlation with only a morphologic facial height.
4) Between indices of head and face, and dental arch and palate, morphologic facial index have heigher correlations with indices of arch and palate than cephalic indices have.

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Primary cilia in the basal cells of main excretory ducts of rat submandibular glands were observed by transmission electron microscope.
About 58% of basal cells had primary cilia which projected in randam directions. The longest of all cilia in the basal cells was about 5μm in lenght. The basal body, which was composed of nine sets triplet microtubules and was accompanied by another centriole, showed a “two centriole type”.
In the most proximal portion of a cilium, doublet microtubules showed a 9+0, while in more distal portion, doublet ones decreased in number and their pattern changed. It seemed that cilia could not move in the absence of central microtubules. Those structures were similar to those of sensory or chemoreceptive cilia. Therefore, it was suggested that the primary cilia in basal cells might be sensory or chemoreceptive.
A primary cilium originated from the depression of basal cell. In more distal portion of cilium, a half desmosome was present between the ciliary membrane and a surrounding basal cell surface.
A few rootlet fibers were present around the basal body of a cilium. Therefore, the basal body may be moved by intracellular movement.
From above observations, it appeared that the tip of primary cilia might be moved passively by basal body movement on the half desmosome as the fulcrum. So, it was suggested that cilia might be engaged in mixing-up of flowing substance in the intercellular space, and might be sensory or chemoreceptive.

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For the purpose of sexing, the discriminant analysis was applied to the measurements of dental arch and palate of Yami, Bunun and Ami, who belong to Formosan aborigines. The results may be summarized as follows.
In the analysis of each tribe, both the discriminant effects derived from 4 measurements of posterior arch and from 4 measurements of anterior arch were not very high. However, the rather high effects of 72.41-77.14% were obtained from 9 measurements of posterior and anterior arch, and palate.
The effects derived from 4 measurements of anterior arch and from 9 measurem ents of arch and palate in the analysis of the combination of three tribes, were considerably lower than in the analysis of each tribe.
A stepwise method employing Wilks' lambda selected 3-5 measurements from 9 measurements of arch and palate, and gave the effects of 67.82-74.51%. The relatively high effects were obtained from the few measurements, though the selected measurements differed among different tribes.
The sexing effect of dental arch and palate was suggested to be nearly as high as crania or teeth, but not so high as appendicular skeletons.

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The fine structure of the lizard (Anolis carolinensis) labial gland was studied by scanning and transmission electron microscopies. The obtained results are as follows.
1) The labial gland of Anolis carolinensis is formed by aggregations of small glands, which can be described as branched tubular and acinar type. The duct of each small gland opens into the vesibulum oris.
2) The glandular cells are arranged in a layer against the basal lamina and appear to be irregulary cuboidal or columnar in shape. The nucleus is at the base of each cell and usually appears ungular. The rough endoplasmic reticulum and the free ribosome are abundant, and are usually located near the basal part of cell.
3) Numerous mucous droplets are contained between the nucleus and the free cell surface, measuring from 1 to 2μm in diameter. Low and high electron dense droplets can be found. Low electron dense droplets are usually irregular in shape and the boundaries between neighbourring droplets are either not clear or completely absent. These mucous droplets seem to be discharged by exocytosis.
4) Another type of cells, the myoepithelial-like cells, containing the droplets similar to the mucous droplets of the glandular cells, were observed among the true myoepithelial cells.
These findings suggest that the fine structure of Anolis carolinensis labial gland is similar to that of human labial gland, inspite of presence of the myoepithelial-like cells.

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The forming enamel surfaces of deciduous molar teeth germs of human fetuses aged from 7 to 9 months were observed by a scanning electron microscopy.
Smooth calcified surfaces and circle-like pits of various diameter (3-5 μm) were seen at the cervical area of the enamel.
The shapes of the pits were divided into three types at the area between the cuspal summit and the cervical area of the enamel. These are; Type I: hexagonal shape (diameter: 5-6 μm), Type II: longitudinal spindle shape (major axis: about 6 μm, minor axis: about 4μm), Type III: mixed type of ovoid, spindle, and rod shape etc.(major axis: 2-6 μm, minor axis: 1-5 μm). Furthermore, a relationship between the arrangement and the shape of these pits was established. The arrangement pattern of the hexagonal and the spindle pits were respectively type HI and type II as described by Boyde, and the irregular pattern was observed in the mixed type. Longitudinal partition (l. p.) which bounded the right and the left pits was well developed but transverse partition (t. p.) which bounded superior and inferior pits did not form well. The concave surface of a pit was composed of two faces, a flat surface (S face) which formed an enamel rod and a curved surface (N face) at which Tomes' process slided.
These two faces were cuspally and cervically directed respectively.
There were few shallow grooves at the cuspal summit area. The pits composing them assumed ellipsoidal shape and had relatively large diameters (6-7μm). The cavity of pit was shallow because of improper development of l.p. and t.p. Consequently, it was not easy to distinguish S from N face.
The relationship between the morphology of enamel surface and the enamel formation was explained based upon these findings.

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Inflammation is a mode of biological defense against noxious agents and proceeds from the stage of tissue damage to the stage of wound healing. In order to obtain an adequate view of the inflammatory process it is important to measure lipid peroxide as being related to tissue damage: indeed, lipid peroxide is an important parameter in the study of inflammation. In connection with the correlation between tissue damage and lipid peroxide in the cell membrane, it is known that the presence of ionic iron in tissue causes an increased production of lipid peroxide.
This paper deals with the impact of ionic iron on the inflamed tissue induced by the dorsumsubcutaneous injection of carrageenin in rats.
Male Wistar rats were divided into three groups: The first group (4 %) received dorsum-subcutaneous injections of 4 % carrageenin solution; the second (2%), 2% carrageenin; the third (Fe), 2 % carrageenin containing FeCl₃ equivalent to 20 mg of Fe (80 mg of Fe caused necrosis). The rats were killed on day 7 or 14 after carrageenin injection.
The Fe and 4% group showed high values compared with the 2 % group in the following substances-LPO, LDH, leucine aminopeptidase (LAP), β-glucuronidase (β-Gl) and fibrin degradatin products (FDP)(all in the pouch fluid), β-Gl and γ-GTP (in the pouch tissue). In the Fe group, tissue weight was lowest with the highest amount of tissue total protein on day 7 (the 4% group showed the highest), while fluid FDP showed the highest values with the lowest tissue total protein on day 14.
The conclusion is that: In carrageenin-induced inflammation ionic iron causes enhancement of the inflammatory response without efficient removal of carrageenin (high LDH, LAP, β-Gl) and degradation of fibrin (high FDP), inhibiting the formation of the granuloma which should grow proportionately to the amount of carrageenin added. The above responses are probably due to excessive production of lipid peroxide (high LPO) caused by the exposure to ionic iron.

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An injection of 25μEl of 0.5% formalin was made into the right hindpaws of mice. The times spent in licking or biting response were measured as indicators of pain response. The response curve was biphasic, having two peaks, from 0 to 5 min (first phase) and from 15 to 20 min (second phase). Using this method, the effects of various analgesic and anti-inflammatory drugs were assessed. Furthermore, the properties of this biphasic pattern and the mechanism of this pain response were investigated and the following results were obtained: 1) Centrally acting drugs such as morphine and pentazocine inhibited pain response at the first phase and the second phase equally. Naloxone and levallorphan reversed morphine analgesia completely. 2) Peripherally acting drugs, aspirin and hydrocortisone inhibited only the second phase significantly. Aminopyrine which acts on both central and peripheral sites inhibited both phases, but the inhibition was stronger in the second phase more than in the first phase. 3) From these results, it is suggested that pain response at the first phase is evoked by direct chemical stimulation of the nerve endings and the second phase is due to inflammatory reaction. Furthermore, this method enables to distinguish roughly the action site of analgesics between the central nervous system and the periphery. 4) The first phase was inhibited by capsaicin pretreatment for sciatic nerve and (D-Pro², D-Trp^{7, 9}) substance P (substance P antagonist) treatment. 5) Compound 48/80 pretreatment inhibited only the second phase significantly. 6) Des-Arg⁹-(Leu⁸)-bradykinin, a specific inhibitor of bradykinin, inhibited both first and second phases. 7) It was recognized that there was a higher correlation between the pain response and vascular permeability especially at the first phase. It is believed that this pain model is useful as a screening method not only for a assessing analgesic effect but for a explanation of the mechanism of analgesic action and for a classification according to the site of action as well.

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Characteristics of the collagenolytic enzyme produced by the pathogen yeast Candida albicans ATCC 1002 were investigated. An enzyme capable of degrading dentine was isolated from the supernate when C. albicans was cultivated in a Yeast Carbon Base containing demineralized dentine.
The enzyme was purified by DEAE-Sephacel chromatography at pH 6.7. The enzyme was shown to be a single substance by means of 7.5% polyacrylamide disc gel electrophoresis.
The molecular weight was found to be 46, 000 by 10 % SDS polyacrylamide gel electrophoresis, the isoelectric point was at pH 4.2.
The optimum pH for collagen degradation lay between 3.5 and 4.0. The enzyme was heat labile and underwent alkaline denaturation in buffer over pH 7.0.
It was observed that the demineralized dentine was degraded by the enzyme at pH 4.0, and a bydroxyproline was detected in the hydrolysate. The enzyme also degraded Azocoll used as a substrate.

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The relationship between functional maturation and sex differences in rat submandibular gland during (2-10 weeks of age) was studied. The secretory properties stimulated by the specific α₁-adrenergic agent, methoxamine, were compared with those by pilocarpine.
The volume of total saliva (during 60 min) was independent of sex, whereas it correlated with the wet weight of the gland. Characteristic differences appeared in the flow rate of saliva, and the concentration of saliva proteins and inorganic ions during development and between the sexes. A positive correlation was observed between saliva Ca²⁺ and protein concentrations in the postnatal changes irrespective of sex differences. This finding suggested that the stimulation of α₁-adrenergic receptors and the release of Ca²⁺ might be correlated with the secretion of saliva proteins. In the comparison of the disk gel electrophoresis profiles of saliva proteins, characteristic changes were observed according to the growth, although not in sex differences. These changes were clearly distinguishable from the response to parasympathomimetics.
The effects of α₁-adrenergic antagonists on the stimulation of the α₁-adrenergic receptors were observed from 2 weeks of age. The gland cyclic-GMP content reached the adult level in 3-4 week old rats, but lowered temporarily in 5 week old ones, and then recovered. Only in 2 week old rats was the increase of cyclic-GMP content corresponding with the stimulation seen, and the response to isoproterenol was also observed in the same age. On the other hand, the increase of cyclic-AMP was seen in 3 week old rats, corresponding with both α- and β-adrenergic receptor stimulations. These results indicate that the various functional changes in rat submandibular gland are correlated with its celldifferentiation.

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Fullmer's oxytalan fibers appear to be special connective tissue fibers belonging to elasticsystem fibers and can be histochemically identified in periodontal ligaments (and some non-dentalsites of certain animal species) with aldehyde fuchsin (AF), orcein, and resorcin fuchsin after peraceticacid or Oxone oxidation. However, it is well known that these methods also stain elastin. We havehistochemically investigated the localization of sulfate groups in oxytalan fibers in monkey (Macaca fuscata) periodontal ligaments, using the high iron diamine (HID), Alcian blue (AB, pH 1.0), AB (pH5.8) +0.65 M MgCl₂, safranin O, and AF with Oxone or periodic acid (PA) oxidation. After Oxoneoxidation oxytalan fibers intensely stained with HID and AF, weakly stained with AB (pH 1.0) and AB (pH 5.8) +0.65 M MgCl₂, and very weakly stained with safranin 0, whereas no staining was observedin oxytalan fibers without Oxone oxidation. These methods except AF did not stain elastin inthe monkey aortic tunica media with or without Oxone oxidation. After PA oxidation oxytalan fibersalso stained weakly to moderately with HID and AF, although this reactivity was not preventedby aldehyde blockage. Thus these studies demonstrate that oxidized oxytalan fibers in monkey periodontalligaments contain sulfate groups and the HID method is a useful tool in studies of oxytalan fibers.

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Formocresol is widely used in endodontic therapy as a root canal disinfective and pulp tissue fixative. Recently, it has been mentioned that autologous soluble components of the dental pulp which are modified by the agent can be rendered antigenically active to the host. Several investigators suggest that such alternative may be due to modification of the components by cresol.
On the other hand, the clinical effect of formalin consists primarily in the action to many sorts of proteins in the root canal system. It is a well known fact that cross-links between protein end-groups are formed when formalin acts on proteins. Cross-linking of proteins may prevent hapten reaching susceptible group.
This study was performed to estimate the inhibitory effect of formalin on the hapten modi - fication of proteins.
Sodium dinitrobenzene sulfonate was used for haptenation of bovine r-globulin (BGG). A product containing about 50 dinitrophenyl groups per molecule of BGG is normally yielded by this procedure.
A lower degree of modification was accomplished when BGG had been treated with formalin.
It was confirmed that formalin had an inhibitory effect on the conjugation of dinitrophenyl groups to BGG.

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To measure oxygen-sensitive pyruvate formate-lyase (EC 2.3.1.54) activity, a spectrophotometerwas placed in an anaerobic glove box (90% N₂, 10% H₂). The activity of pyruvateformate-lyase of Streptococcus mutans strain NCIB 11723 obtained by this method was at least eightfoldhigher than that obtained by the Thunberg tube method. This assay method is considered moreaccurate and more sensitive, and it allows a continuous assay of the enzyme activity when lactatedehydrogenase activity is absent in the assay mixture. When pyruvate formate-lyase was incubatedwith a low concentration of oxygen, the activity was decreased with time. At a fixed concentrationof oxygen the lower the concentration of the enzyme was, the more activity was lost. The inactivationby oxygen was ascribed to the production of neither superoxide nor hydrogen peroxide butmolecular oxygen itself. It was suggested that this method could be used for the accurate assay ofnot only pyruvate formate-lyase but also other oxygen-sensitive enzymes.

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Scanning electron microscopy was employed to investigate the ultrastructure of oral epithelialcells of the lizard, Takydromus tachydromoides. The specimens were prepared by a methodinvolving osmium postfixation and acid treatment to remove extracellular attached material.
Bump-shaped protuberances were arranged symmetrically on both sides of the median line of thepalate. Epithelial surfaces besides the bump-shaped protuberances were widely covered with fine folds.At higher magnification, a network pattern of microridges was closely distributed over the entiresurface of the palate, and each cell marginal thickening was clear. The protuberances were alsoobserved in the latero-posterior region of the floor of the mouth. A number of nicks were arrangedsymmetrically on both sides along the median line of the fore-region of the floor. Taste buds werescattered in the epithelial surfaces between nicks. At higher magnification, microvilli as well as microridgeswere observed on the epithelial cell surfaces of the floor. There were no papillar structures onthe ventral tongue surface. Fine pits were densely distributed over the entire epithelial cellsurface. The cell margin was clearly distinguishable as a depressed line. The epithelial surface ofthe laryngeal part of the pharynx was entirely covered with ciliated cells. Taste buds were alsoscattered on the inner gingival epithelial surfaces of the upper and lower jaws along the dentalarches. Relatively indistinct microridges were widely distributed on the gingival epithelial cell surfaceof the upper jaw. Scrollwork and network patterns of microridges were clearly observed on the gin -gival epithelial surface of the lower jaw.

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For elucidation of microbial effects on rat lymph node microcirculation, intravascular andintralymphatic cast preparations were examined along with histological observation, using light microscopy.Submandibular lymph nodes from conventional (CV), specific-pathogen-free (SPF), antibioticdecontaminated-SPF (AB-SPF) and germfree (GF) rats were studied.
In nodes exposed to microbial stimulation (CV and SPF), typical high-endothelial-venules (HEVs) with many lymphocytes within their walls, received abundant vascular connections from the vascularplexuses in the superficial cortex and assumed close proximity to the intermediate sinuses plugged withmany lymphocytes. In nodes with almost no microbial stimulation (AB-SPF and GF), overall vascularitywas sparse, and the vascular connections to the HEVs with few lymphocytes in their walls wereminimal. In the AB-SPF rat nodes, the intermediate sinuses were often vacant, and in the GF ratnodes, they were poorly developed. Arteriovenous anastomoses were frequently observed to join HEVs, regardless of differences in microbial conditions.
Therefore, in rat lymph nodes, microbial effects seem to induce a rich blood supply to the HEVsby organizing vascular plexuses in the superficial cortex. This promotes the active traffic of lymphocytesfrom the HEVs into the nodes and formation of intermediate sinuses, and related functions areenhanced.

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Bacteria lytic against heat-killed cells of Streptococcus (S.) sanguis ATCC 10558 were examined in dental plaques of 20 infants. Such bacteria were isolated from dental plaques taken from pits and fissures of the healthy deciduous molars opposed to carious molars of all subjects, but not from those on labial cervical surfaces of sound inciso rs opposed to sound incisors. Biological characterizations have shown that all of these lytic bacteria are S. mutans.

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Lytic ability and bacteriocin activity against Streptococcus (S.) sanguis ATCC 10556, 10557 and 10558 and artificial plaque forming ability by 119 strains of S. mutans were examined. Previously we have shown that 104 strains were serotype c/e/f and 15 were d/g.
Serotype c/e/f strains showed a fairly good lytic ability against S. sanguis ATCC 10558, but not against ATCC 10556 and 10557. They showed a higher bacteriocin activity against S. sanguis ATCC 10556 and 10557 than against ATCC 10558. In serotype d/g strains, a slight lytic ability against S. sanguis ATCC 10558 but not two others was seen, and an extremely slight bacteriocin activity against all S. sanguis strains was noted. The artificial plaque forming ability by d/g type strains was clearly higher than c/e/f type strains. These facts seem to show that these factors from S. mutans compensate for the defects of each other. From these results, it is probable that not only bacteriocin activity, but the lytic ability and artificial plaque forming ability of S. mutans also take part in the decrease of S. sanguis in the dental plaque.

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