Immunohistochemical localization of glycosaminoglycans and proteoglycans in predentin and dentin of rat incisors.

We examined immunocytochemically the type and distribution of glycosaminoglycans and proteoglycans (PG) in predentin and dentin demineralized with EDTA after aldehyde fixation of rat incisors using (a) four monoclonal antibodies (1-B-5,9-A-2,3-B-3, and 5-D-4) which recognize epitopes in unsulfated chondroitin (C0-S), chondroitin 4-sulfate (C4-S), chondroitin 6-sulfate (C6-S), and keratan sulfate (KS) associated with the PG, and (b) monoclonal (5-D-5) and polyclonal antibodies specific for the core protein of large and small dermatan sulfate (DS) PG. Light microscope immunoperoxidase staining after pre-treatment of tissue sections with chondroitinase ABC localized the majority of stainable PG (C4-S, KS, DSPG, C0-S, and C6-S) in predentin and, to a lesser extent (C4-S and small DSPG), in the dentin matrix. The former site demonstrated relatively homogeneous PG distribution, whereas the latter site revealed that strong staining of C4-S and small DSPG was confined mostly to dentinal tubules surrounding odontoblastic processes, with only weak staining in the rest of the dentin matrix. These results indicate that there is not only a definite difference between PG of predentin and dentin but also a selective decrease in the concentration or alteration of these macromolecules during dentinogenesis and mineralization.

Neuropeptides in the dental pulp: distribution, origins, and correlation.

It is known that many factors participate in the process of inflammation and pain transmission. Recently, various neuropeptides have been demonstrated in the intrapulpal nerve fibers immunobiohistochemically. These neuropeptides may play some role in the process of inflammation and transmission of pain sensation. In this presentation, the distribution, origins, and correlation of neuropeptides in the dental pulp is reviewed.

Neuropeptide K-like immunoreactivity in human dental pulp.

Nerve fibres displaying such immunoreactivity were revealed by indirect immunofluorescence. Neuropeptide K-like immunoreactive fibres, entering the pulp within large nerve trunks, were distributed around blood vessels as well as in the stroma. Some immunoreactive fibres were also observed in the para-odontoblastic region. In view of the biological activity of neuropeptide K, it is tentatively proposed that it may act in the dental pulp as a regulatory peptide involved in neurogenic inflammation, blood flow regulation and sensory transmission.

Bovine dental pulp collagens: characterization of types III and V collagen.

Pulp was essentially solubilized by partial pepsin digestion. The various genetic types of collagens were isolated by differential salt precipitation and extraction. Types I, III and V collagen represented 56, 41 and 2% of the total collagen, respectively. The type V collagen comprised two different molecular species consisting of [alpha 1(V)]2 alpha 2(V) and alpha 1(V) alpha 2(V) alpha 3(V), the ratio of which was approx. 1:1.3. The major portion of the type III collagen was present as a high molecular weight aggregate which released alpha 1(III) chains upon reduction with 2-mercaptoethanol.

Detection of HIV in the dental pulp of a patient with AIDS.

Dental pulp tissue from a patient with acquired immune deficiency syndrome (AIDS) was examined to determine the presence of the human immunodeficiency virus (HIV). The results found a high concentration of proviral HIV DNA.

Responses of pulpal nerves to cavity preparation in rat molars: an immunohistochemical study using neurofilament protein (NFP) antiserum.

The response of neural elements to a dentin injury was morphologically investigated in rat molars by use of immunostaining for neurofilament protein (NFP). An artificially formed cavity in dentin by drilling rapidly caused the displacement of some odontoblasts into the exposed dentinal tubules, while others were detached from the predentin. The subodontoblastic nerve plexus consisting of NFP-immunoreactive nerves shifted inward together with the separated odontoblasts, while a movement of the nerves into the exposed dentinal tubules was not recognized. The odontoblasts separated from the predentin degenerated and disappeared one day after the cavity preparation; at this time, the subodontoblastic nerve plexus underlying the drilled dentin was remarkably disrupted, presumably losing dentinal sensation of the drilled area. Three days after the cavity preparation, the destroyed odontoblastic layer began to be repaired by newly differentiating odontoblasts; the reparative dentin was produced from 5 to 7 days onward. Numerous NFP-positive nerves, beaded in type, gathered in the odontoblastic layer in accordance with the differentiation of the new odontoblasts. The increased beaded nerve fibers were suggested to represent peptide-containing nerves. In 10-15 days, the reparative dentin accumulated quite remarkably under the cavity area. The NFP-positive subodontoblastic nerve plexus was entirely reconstituted and also regained continuity to its surrounding plexus. The nerve fibers in the reconstituted plexus were mostly non-beaded in type as seen in the control teeth. Since dentinal tubules in the reparative dentin are not normally continuous to the primary dentinal tubules, dentinal sensation may not have been restored.

Simultaneous determination of FUC and PGM1 by isoelectric focusing.

This paper describes a simple procedure for the simultaneous detection of FUC and PGM1 in semen, seminal stains, dental pulps and hair roots after separation by isoelectric focusing using Ampholine pH 5-7. The gel plate was first stained for FUC using an agarose overlay with a buffer of pH 7.5. After reading the FUC types, the FUC overlay was removed and replaced with an agarose overlay for PGM. The results show that the method is useful in medicolegal individualization of seminal stains and teeth when only small samples have to be examined.

Analysis of DNA from tooth and application to forensic dental medicine.

The DNA extracted from teeth was examined to find out the usefulness in forensic dental medicine. Samples of tooth-derived DNA were digested with a restriction enzyme, electrophosed through agarose gel and subjected to Southern blot hybridization, using a 32P-labeled minisatellite DNA probe "Myo" and a Y chromosome-specific DNA probe. The results indicate that DNA extracted from dental pulp is useful for identification of individuals, paternity testing, and sex determination.

Adenine nucleotide contents and ATPases activities in porcine deciduous dental pulp during the root formation, in fully formed root and during root resorption phases.

Nucleotide content and activity of certain enzymes were compared in pigs of various ages in order to study the energetic metabolism of deciduous dental pulps in the three phases of the cycle of tooth ontogeny, namely, root formation, fully formed root and root resorption phases. The frozen pulps were removed with the help of a screw vise and analysed for ATP, ADP and AMP contents and Ca2+ and Mg2+-ATPases activities. The highest ATP content in the first deciduous molar pulp was found when the tooth was still in an intrabony position. The calculated energy charge, although low for all groups, at this stage of development, indicated an activation of the consuming processes. In the root resorption phase, lowest ATP content and higher Ca2+ and Mg2+-ATPases activities were observed.

Effects of orthodontic force on methionine enkephalin and substance P concentrations in human pulpal tissue.

Orthodontic treatment typically involves intermittent periods of patient discomfort caused by forces on the teeth and adjacent tissues. This sensation of discomfort presumably is caused by the action of neuropeptides in the peripheral and central nervous systems. The effects of orthodontic force on the concentrations of two endogenous neuropeptides, methionine enkephalin (ME) and substance P (SP), measured as immunoreactive-methionine enkephalin (ir-ME) and immunoreactive-substance P (ir-SP), in human tooth pulp were evaluated in 20 patients from whom premolars were extracted before orthodontic treatment. The teeth from nine controls were not subjected to a force, whereas the 11 experimental patients had force applied to their maxillary premolars either by a transpalatal spring ligature or, in one case, by a headgear. The ligature applied a force within the range of 120 to 245 gm; the headgear applied 600 gm. Reversed-phase high-performance liquid chromatography (RP-HPLC) was used to purify the neuropeptides in the pulp homogenate, and radioimmunoassay (RIA) was used to quantify ir-ME and ir-SP in their appropriate HPLC fractions. (1) Females subjected to orthodontic force had significantly greater ir-ME concentrations than males. (2) The ir-SP concentration decreased significantly from the first to the third tooth extracted, then increased from the third to the fourth tooth. (3) Ir-SP and ir-ME concentrations are positively intercorrelated. The association was highest in the first tooth extracted from controls; surgical extraction decreased the correlation, although it continued to be positive. (4) The concentrations of ir-ME and ir-SP each correlated negatively with the magnitude of the orthodontic force and that correlation was enhanced when the value of the force was log-transformed.

[Peptidergic innervation of the human dental pulp: neuropeptide Y]. / Innervazione peptidergica della polpa dentaria umana: il neuropeptide Y.

The localization and distribution of the Neuropeptide Y (NPY) has been studied with immunofluorescent methods in the human dental pulp. Immunofluorescence for the NPY has been observed in nervous fibers running in medium and big nerves associated in vascular structures, and in single fibers scattered in the pulpar connective or organized in the subdontoblastic plexus.

Autometallography for histochemical visualization of rat incisor polyanions with cuprolinic blue.

Autometallography was applied to semi-thin sections of rat incisors fixed a solution of cuprolinic blue-aldehyde. The resulting reduction of silver ions to metallic silver amplifies the copper sulfide signal of the cationic dye. Silver grains were seen over the cell bodies of ameloblasts and odontoblasts but not over their processes. This was owing to the interaction of cuprolinic blue with the DNA and RNA of these cells. In the extracellular matrix, silver grains were unevenly distributed over the predentin, dentin, and forming enamel. The distal predentin near the mineralization front and a thin band of dentin located near the dentino-enamel junction displayed unexpectedly intense accumulation of silver grains, whereas all other portions of the extracellular matrix exhibited the distribution of glycosaminoglycans expected from previous studies. The present investigation constitutes a new application of autometallography to glycosaminoglycan histochemistry.

[Studies on sex determination using human dental pulp. II. Sex determination of teeth left in a room].

The appearance rate of Y chromatin was calculated in the dental pulp with the staining by Quinacrine Mustard. The morphological findings of the tissue were observed with the HE stained section and the change of the appearance rate of Y chromatin was studied along with the morphological change of the tissue. The average of the appearance rate of Y chromatin in the male dental pulp was 42.2% immediately after the tooth extraction, 34.8% after one month, 27.9% after 3 months, 20.8% after 6 months, 20.6% after 1 year, 19.1% after 18 months and 11.4% after 2 years, respectively. On the other hand, the appearance rate of the pseudo-Y chromatin spot in the female pulp was below 4.2% through the two years' observation which enabled to determine the sexuality. By the HE staining observation about the dental pulp, left at a room temperature, it was revealed that the pulp tissue can be well preserved for a long time because of the natural drying. The degree of the staining declined with the lapse of time after the removal of tooth. Degeneration of tissue did not offer any discrete criteria for the estimation of the lapse of time after the removal of tooth.

Immunoreactive evidence of beta-endorphin and methionine-enkephalin-Arg-Gly-Leu in human tooth pulp.

For the first time, beta-endorphin-like immunoreactivity (BE-LI) has been measured in human tooth pulp. Separation of peptides from pulp tissue was achieved by acid extraction followed by chromatographic separation through a Sep-Pak disposable cartridge. Reversed phase high performance liquid chromatographic (RP-HPLC) was performed on the peptide-rich fractions for further peptide separation. Radioreceptor assay (RRA) data of the HPLC fractions was used to construct a profile of opioid-receptor active peptides. Radioimmunoassay (RIA) data provided further information. Following acute mechanical stress, a monotonic decrease in BE-LI concentrations was evident according to a four bicuspid extraction order.

[The significance of endogenous peptides for dental pulp pain]. / Zur Bedeutung der endogenen Peptide für den pulpalen Schmerz.

The increase of knowledges in the field of endorphines and other peptides causes new aspects of development and transmission of dental pain. Methionin-enkephalin (ME) and substance P (SP) are found also in the dental pulp and the dentine. The concentration of SP is higher in the pulp, than in some other tissues. The concentration of both, ME and SP, in the endodont is different with respect to different functional situations, e.g. it is lower in case of pain. We suppose, there is a peptiderg reception and transmission of pain in dentine. Interaction with this new system opens new ways for opposing pulpal pain.

Comparative study of the composition of primary and secondary dentine.

There is considerable variation in the composition of primary and secondary dentine. In general, secondary deposits, regardless of type, contain less calcium, phosphorous, and collagenous matrix per unit volume when compared to primary dentine. Fluoride levels show the reverse tendency. These observations suggest a more open, porotic structure for secondary deposits.

S-100 protein localization in human adult dental pulp.

The pulp of third molar teeth was examined by immunofluorescence and immunoperoxidase using a specific antibody to detect S-100 protein-labelled cells. There was a strong positive reaction in macrophages and in Schwann cells ensheathing axons. The other cells within the pulp were negative.

Aspiration and characterization of predentin fluid in developing rat teeth by means of a micropuncture and micro-analytical technique.

A fluid phase was aspirated in vivo and in vitro from predentin or pulp of developing rat teeth by means of a micropuncture technique. Pooled aspirates (approx. 2 nL) were analyzed for P, Na, K, Ca, Mg, and S by electron probe microtechniques (Lechene and Warner, 1979). Compared with pulp fluid, currently and previously studied cartilage fluids, as well as serum, predentin fluid showed elevated K, depressed Na, Cl, and Ca, as well as increased P. Statistical analysis was possible for only a few groups of comparisons among the elemental profiles. Ultrastructural examination of the aspiration site and of the aspirates showed no evidence of contamination with cell organelles or other formed elements. The micropuncture technique used was a critically precise and laborious procedure; possible contamination with intracellular fluid could not be avoided. The consistently low Mg concentration found in the aspirates, however, supports our view that the samples were primarily extracellular.