Scanning electron microscopic study of odontoblasts and circumpulpal dentin in a human tooth Dominique Marion,O Alain Jean,a Henri Hamel,b Lise-Marie Bertrand Kerebel,d Nantes, France INSERM
RESEARCH
UNIT
AND
DENTAL
Kerebel, PhD,C and
FACULTY
Two combined scanning electron microscopic methods including modified fixative procedures were used for studying the morphologic aspects of the odontoblasts and the corresponding dentinal wall at different endodontic levels. The odontoblasts were tightly packed in the pulp horn, where they assumed pear-shaped profiles and, from crown to apex, successively looked spindle shaped, club shaped, and globular. Their number decreased in the pulp radicular portion, where the interglobular spaces were enlarged. The filling fibrillar material varied from crown to apex. A globular circumpulpal dentin was observed at all levels of the root canal. The tubule openings varied in number and size. (ORAL SURG ORAL MED T)RAL PATHOL
1991;72:473-8)
I
n the last decadescanningelectron microscopy @EM) hasbeenusedto checkthe efficiencyof endodonticinstrumentsandof varioussolutionsusedfor canalirrigation.l-4 However,the variousparietalareasof the endodonticsystemarenot yet well known. Becauseof technicaldifficultiesrelatedto fixationand preparationprocedures,reliable SEM observations are uncommon.5Actually, surfaceconditionsafter instrumentalproceduresare describedwithout referenceto the initial state.The purposeof the present studywasto developa descriptivecatalogof the morphologicaspectsof the odontoblastsand the correspondingdentinalwall at differentendodonticlevels with two combinedSEM methods. MATERIAL
AND METHODS
A soundupperleft first premolarwasremovedsurgically from a 43-year-oldwoman whoseocclusion BProfessor, Department of Endodontics. bProfessor and Chief, Department of Endodontics, and Dean of Dental Faculty. CResearcher. dProfessor and Chief, Department of Biology and Basic Sciences, and Director, INSERM Unit U. 225. 7/15/28788
Fig. 1. Overall view of odontoblasts. magnification, X370.)
(SEM;
original
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ORAL
SURG ORAL
MED
ORAL
PATHOL
October 1991
Fig. 2. Overall view of circumpulpal dentin. @EM; original magnification, X370.)
Fig. 4. Aspect of dentin in pulp horn. (SEM; original magnification, X2300.)
Fig. 3. Pulp horn. Note pear-shaped aspect of odontoblasts. (SEM; original magnification, X2300.)
Fig. 5. Spindle-shaped aspect of odontoblasts at proxima1midpulpal level. (SEM; original magnification, X2300.)
Volume 72 Number 4
SEM study of human odontoblasts and dentin
475
6. Aspect of dentin at root coronal third level. @EM; original magnification, x2300.)
Fig.
6. Aspect of dentin at proximal midpulpal level. (SEM; original magnification, X2300.)
Fig.
Fig. 7. Odontoblasts assuming club-shaped profiles at root coronal third level. (SEM; original magnification, x2300.)
Fig.
9. Odontoblasts assuming short, club-shaped profiles at root midthird level. (SEM; original magnification, x2300.)
476
Marion et al.
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SURG
ORAL
MED
ORAL
PATHOL
October 199I
Fig. 10. Note smooth aspect of globular dentinal relief at root midthird level. (SEM; original magnification, X2300.)
Fig.
was normal. Extraction was performed with the patient under local anesthesia without epinephrine. The tooth was fractured along its longitudinal axis, and the specimens were prepared for SEM examination according to a previously described method5 involving postfixation in tannic acid. The cytologic aspects of the odontoblasts at different levels of the pulp chamber and radicular canal were studied with Hitachi S 450 and JEOL JSM 35 scanning electron microscopes. The same specimens were then cleaned ultrasonically for 5 minutes in a 2.5% sodium hypochlorite solution, prepared again for SEM, and examined with both aforementioned microscopes, to study the mineral aspects of the circumpulpal dentin corresponding to the areas where the odontoblasts had been observed previously.
varied in length, seemingly assuming a stratified aspect. The underlying dentin (Fig. 4) was pitted with several wide tubules (2 pm in diameter) and had flattened profiles. The disjoined dentinal globules were irregular in shape and often notched, and varied in size. At the proximal midpulpal level the spindle-shaped odontoblasts (Fig. 5) looked less tightly packed than described earlier. The intercellular spaces were filled with fibrillar structures with spiral profiles. The surface of the corresponding dentinal wall (Fig. 6) appeared evenly globular. The globules were joined and round. Only a few of them had the notched aspect. There were as many tubule openings as previously observed, and their diameter was about the same. At the root coronal third level the cell bodies (Fig. 7) were randomly distributed, with enlarged intercellular spaces containing a scanty fibrillar material. They assumed club-shaped profiles, and their bulging pulpal pole continued in a short pedicle. The dentinal globules (Fig. 8) were variable in size, and less tightly packed than previously observed, inducing interglobular spaces. The tubule openings were reduced in number, randomly distributed, and smaller in diameter (1.5 to 2 pm). At the root midthird the intercellular spaces were enlarged (Fig. 9) and filled with an ill-defined fibrillar
RESULTS
The overall views of the same area of the circumpulpal dentin examined twice showed first the space distribution of the odontoblasts (Fig. l), then the globular aspect of the dentin after removal of the odontoblasts (Fig. 2). At the pulp horn level the odontoblastic cell bodies appeared as elongated pears (Fig. 3) arranged in close contact with each other. The basal (pulpal) pole was bulging. The cell bodies were 5 pm in diameter and
11. Odontoblasts seen as disjoined globules at root apical third level. (SEM; original magnification, X2300.)
Volume 72 Number 4
SEM study of human odontoblasts and dentin
477
Fig. 13. Diagram summarizing shape variations of odontoblasts. a, Pulp horn (pear shape); b, proximal midpulpal level (spindle shape); c, root coronal third level (elongated club shape); d, root midthird (short club shape); e, root apical third (globules).
12. Aspect of dentin at root apical third level. @EM; original magnification, X2300.)
Fig.
material. The cell bodies assumed short, clubbed profiles. The whole cell body, not only the basal pole, was bulging. The circumpulpal dentin (Fig. 10) showed a smoother globular relief. The tubule openings were fewer and varied in diameter (1.5 to 2 pm) as in the coronal third. At the root apical third the cell bodies (Fig. 11) were definitely disjoined and assumed compact globular profiles. The intercellular spaces were filled with profuse fibrillar material arranged in large bundles or in fine fibrils. The dentinal wall (Fig. 12) had globules with a smooth surface. The scanty tubules were randomly distributed and small in diameter (1 pm); some were in the process of obliteration. The various shapes of the odontoblasts are summarized in Fig. 13. DISCUSSION
The study of the functional unit constituted by the pulpodentinal complex is not easy, because of the interface between a mineralized structure and highly differentiated cells. Histologic examination provides only single-plane views of the odontoblasts, and until recently’ SEM images showed many artifacts. Our technique allows to reduce SEM artifacts as shown by measurements of the cell processes at the basal pole,
which correspond exactly to those of the tubule openings. It is therefore possible to compare the results obtained from SEM examination of both the coronal and radicular zone, which has not yet been performed. The present SEM survey along the whole circumpulpal dentin shows that the odontoblasts form a single layer, which often assumes a pseudostratified aspect when examined by light microscopy.6“4 The cell bodies are arranged vertical to the dentinal surface. When measured at the coronal level, their size is similar to that reported by other investigators 799, 15-17 Classic data generally refer to the dentinal coronal level. A change in the structure of the odontoblasts, columnar in the crown and more cuboid in the apical area, has been reported on the basis of histologic sections only.16y I7 The present investigation provides assessment of the three-dimensional aspects of the odontoblasts at different endodontic levels. They appear tightly packed in the pulp horn and successively pear shaped, spindle shaped, club shaped, or globular from the crown to the apex. The intercellular spaces observed might be due to a decrease in the number of cells with age.16 Considering our patient’s age (43 years), it is obvious that dentinogenesis is in a less active synthetic state than during the first stages of dentinogenesis. The disjoined cells described in the present study might be the result of a disruption of the intercellular connections reported in transmission electron microscopy10 and SEM,S which means that the odontoblasts might be in a resting state.17 The surface conditions of the circumpulpal dentin have been studied at the coronal level only, during the active stage of dentinogenesis.18 The three-dimensional variations in the structure of the dentinal surface at the coronal and radicular levels have never been compared before. The present data show the dentinal surface to be evenly globular, whereas this
478
Marion et al.
ORAL
7. 8. 9. 10. 11. 12. 13.
REFERENCES 1. Goldman LB, Goldman M, Kronman JH, Lin P.S. Scanning electron microscope study of a new method in endodontic treatment. ORAL SURG ORAL MED ORAL PATHOL 1979;48: 79-83. 2. Cunningham WT, Martin H. A scanning electron microscope evaluation of root canal debridement with the endosonic ultrasonic synergetic system. ORAL SURG ORAL MED ORAL PATHOL 1982;53:527-31.
3. Auther A, Laurichesse JM, Launay Y, Calas P. Etude au microscope a balayage des surfaces radiculaires traittes par les instruments ultrasonores. Rev Fr Endod 1984;3:29-39. 4. Dietschi JM, Ciucchi B, Cergneux M, Holz J, Baume LJ. Preparation du canal radiculaire a I’aide d’ultrasons: etude au MEB. Rev Fr Endod 1984;3:15-27. 5. Jean A, Kerebel B, Kerebel LM. Scanning electron microscope study of the predentin-pulpal border zone in human dentin. SURG ORAL
MED ORAL
MED ORAL
PATHOL
October 1991
type of mineralization has been reported to occur only occasionally along the predentinal forming front.9 The main variations involve the size and shape of the calcospherites, and the number and size of the tubule openings. On the basis of the precise data provided by the present study on the structure of the odontoblasts and of the dentinal surface at the different endodontic levels, it should soon be possible to corroborate the morphologic variations with the cytophysiologic data provided by other techniques and thus acquire a better knowledge of the endodontic functional unit.
ORAL
SURG ORAL
PATHOL
1986;61:392-8.
6. Zerosi C. Compandio di odontoiatrica conservativa terapia,
14. 15. 16. 17. 18.
Ricostruttiva e Endodontia. Milan, Italy: Scienza e Tecnica dentistica, Edizioni Internazionali. 1982:8-26. Seltzer S. Bender IB. The dental .oul~.. 3rd ed. Philadeluhia: JB Lippincott, 1983. Mjor IA, Fejerskov 0. Histology of the human tooth. 3rd ed. Copenhagen, Denmark: 1985, Munskgaard. Bhaskar SN. Orban’s oral histology and embryology. 10th ed. St Louis: CV Mosby, 1986. Frank RM. Etudeau microscopeelectroniquede I’odontoblaste et du canalicule dentinaire humain. Arch Oral Biol 1966;ll: 179-99. Takuma S, Nagai N. Ultrastructure of rat odontoblasts in various stages of their development and maturation. Arch Oral Biol 1971;16:993-1011. Holland CR. Membrane junctions on cat odontoblasts. Arch Oral Biol 1975;20:55 1-2. Viti M, Masi PL. Ultrastructural aspects of interodontoblastic junctions. Bull Group Int Rech Sci Stomatol Odontol 1981;24:61-9. Sasaki T, Nakagawa K, Higashi S. Ultrastructure of odontoblasts in kitten tooth germs as revealed by freeze-fracture. Arch Oral Biol 1982;27:897-904. Scott JH, Symons NBB. Introduction to dental anatomy. 9th ed. Edinburgh: Churchill Livingstone, 1982:216-48. Ten Cate AR. Oral histology: development, structure and function. 2nd ed. St Louis: CV Mosby, 1985. Provenza DV, Seibel W. Oral histology. 2nd ed. Philadelphia: Lea & Febiger, 1982. Lester RS, Boyde A. Electron microscopy of predentinal surfaces. Calcif Tiss Res 1967; 1:44-54.
Reprint
requests:
Dominique Marion Faculte de Chirurgie Dentaire Pl. A. Ricordeau 44042 Nantes Cedex, France
RESEARCH
UNIT
AND
DENTAL
Kerebel, PhD,C and
FACULTY
Two combined scanning electron microscopic methods including modified fixative procedures were used for studying the morphologic aspects of the odontoblasts and the corresponding dentinal wall at different endodontic levels. The odontoblasts were tightly packed in the pulp horn, where they assumed pear-shaped profiles and, from crown to apex, successively looked spindle shaped, club shaped, and globular. Their number decreased in the pulp radicular portion, where the interglobular spaces were enlarged. The filling fibrillar material varied from crown to apex. A globular circumpulpal dentin was observed at all levels of the root canal. The tubule openings varied in number and size. (ORAL SURG ORAL MED T)RAL PATHOL
1991;72:473-8)
I
n the last decadescanningelectron microscopy @EM) hasbeenusedto checkthe efficiencyof endodonticinstrumentsandof varioussolutionsusedfor canalirrigation.l-4 However,the variousparietalareasof the endodonticsystemarenot yet well known. Becauseof technicaldifficultiesrelatedto fixationand preparationprocedures,reliable SEM observations are uncommon.5Actually, surfaceconditionsafter instrumentalproceduresare describedwithout referenceto the initial state.The purposeof the present studywasto developa descriptivecatalogof the morphologicaspectsof the odontoblastsand the correspondingdentinalwall at differentendodonticlevels with two combinedSEM methods. MATERIAL
AND METHODS
A soundupperleft first premolarwasremovedsurgically from a 43-year-oldwoman whoseocclusion BProfessor, Department of Endodontics. bProfessor and Chief, Department of Endodontics, and Dean of Dental Faculty. CResearcher. dProfessor and Chief, Department of Biology and Basic Sciences, and Director, INSERM Unit U. 225. 7/15/28788
Fig. 1. Overall view of odontoblasts. magnification, X370.)
(SEM;
original
473
474
Marion et al.
ORAL
SURG ORAL
MED
ORAL
PATHOL
October 1991
Fig. 2. Overall view of circumpulpal dentin. @EM; original magnification, X370.)
Fig. 4. Aspect of dentin in pulp horn. (SEM; original magnification, X2300.)
Fig. 3. Pulp horn. Note pear-shaped aspect of odontoblasts. (SEM; original magnification, X2300.)
Fig. 5. Spindle-shaped aspect of odontoblasts at proxima1midpulpal level. (SEM; original magnification, X2300.)
Volume 72 Number 4
SEM study of human odontoblasts and dentin
475
6. Aspect of dentin at root coronal third level. @EM; original magnification, x2300.)
Fig.
6. Aspect of dentin at proximal midpulpal level. (SEM; original magnification, X2300.)
Fig.
Fig. 7. Odontoblasts assuming club-shaped profiles at root coronal third level. (SEM; original magnification, x2300.)
Fig.
9. Odontoblasts assuming short, club-shaped profiles at root midthird level. (SEM; original magnification, x2300.)
476
Marion et al.
ORAL
SURG
ORAL
MED
ORAL
PATHOL
October 199I
Fig. 10. Note smooth aspect of globular dentinal relief at root midthird level. (SEM; original magnification, X2300.)
Fig.
was normal. Extraction was performed with the patient under local anesthesia without epinephrine. The tooth was fractured along its longitudinal axis, and the specimens were prepared for SEM examination according to a previously described method5 involving postfixation in tannic acid. The cytologic aspects of the odontoblasts at different levels of the pulp chamber and radicular canal were studied with Hitachi S 450 and JEOL JSM 35 scanning electron microscopes. The same specimens were then cleaned ultrasonically for 5 minutes in a 2.5% sodium hypochlorite solution, prepared again for SEM, and examined with both aforementioned microscopes, to study the mineral aspects of the circumpulpal dentin corresponding to the areas where the odontoblasts had been observed previously.
varied in length, seemingly assuming a stratified aspect. The underlying dentin (Fig. 4) was pitted with several wide tubules (2 pm in diameter) and had flattened profiles. The disjoined dentinal globules were irregular in shape and often notched, and varied in size. At the proximal midpulpal level the spindle-shaped odontoblasts (Fig. 5) looked less tightly packed than described earlier. The intercellular spaces were filled with fibrillar structures with spiral profiles. The surface of the corresponding dentinal wall (Fig. 6) appeared evenly globular. The globules were joined and round. Only a few of them had the notched aspect. There were as many tubule openings as previously observed, and their diameter was about the same. At the root coronal third level the cell bodies (Fig. 7) were randomly distributed, with enlarged intercellular spaces containing a scanty fibrillar material. They assumed club-shaped profiles, and their bulging pulpal pole continued in a short pedicle. The dentinal globules (Fig. 8) were variable in size, and less tightly packed than previously observed, inducing interglobular spaces. The tubule openings were reduced in number, randomly distributed, and smaller in diameter (1.5 to 2 pm). At the root midthird the intercellular spaces were enlarged (Fig. 9) and filled with an ill-defined fibrillar
RESULTS
The overall views of the same area of the circumpulpal dentin examined twice showed first the space distribution of the odontoblasts (Fig. l), then the globular aspect of the dentin after removal of the odontoblasts (Fig. 2). At the pulp horn level the odontoblastic cell bodies appeared as elongated pears (Fig. 3) arranged in close contact with each other. The basal (pulpal) pole was bulging. The cell bodies were 5 pm in diameter and
11. Odontoblasts seen as disjoined globules at root apical third level. (SEM; original magnification, X2300.)
Volume 72 Number 4
SEM study of human odontoblasts and dentin
477
Fig. 13. Diagram summarizing shape variations of odontoblasts. a, Pulp horn (pear shape); b, proximal midpulpal level (spindle shape); c, root coronal third level (elongated club shape); d, root midthird (short club shape); e, root apical third (globules).
12. Aspect of dentin at root apical third level. @EM; original magnification, X2300.)
Fig.
material. The cell bodies assumed short, clubbed profiles. The whole cell body, not only the basal pole, was bulging. The circumpulpal dentin (Fig. 10) showed a smoother globular relief. The tubule openings were fewer and varied in diameter (1.5 to 2 pm) as in the coronal third. At the root apical third the cell bodies (Fig. 11) were definitely disjoined and assumed compact globular profiles. The intercellular spaces were filled with profuse fibrillar material arranged in large bundles or in fine fibrils. The dentinal wall (Fig. 12) had globules with a smooth surface. The scanty tubules were randomly distributed and small in diameter (1 pm); some were in the process of obliteration. The various shapes of the odontoblasts are summarized in Fig. 13. DISCUSSION
The study of the functional unit constituted by the pulpodentinal complex is not easy, because of the interface between a mineralized structure and highly differentiated cells. Histologic examination provides only single-plane views of the odontoblasts, and until recently’ SEM images showed many artifacts. Our technique allows to reduce SEM artifacts as shown by measurements of the cell processes at the basal pole,
which correspond exactly to those of the tubule openings. It is therefore possible to compare the results obtained from SEM examination of both the coronal and radicular zone, which has not yet been performed. The present SEM survey along the whole circumpulpal dentin shows that the odontoblasts form a single layer, which often assumes a pseudostratified aspect when examined by light microscopy.6“4 The cell bodies are arranged vertical to the dentinal surface. When measured at the coronal level, their size is similar to that reported by other investigators 799, 15-17 Classic data generally refer to the dentinal coronal level. A change in the structure of the odontoblasts, columnar in the crown and more cuboid in the apical area, has been reported on the basis of histologic sections only.16y I7 The present investigation provides assessment of the three-dimensional aspects of the odontoblasts at different endodontic levels. They appear tightly packed in the pulp horn and successively pear shaped, spindle shaped, club shaped, or globular from the crown to the apex. The intercellular spaces observed might be due to a decrease in the number of cells with age.16 Considering our patient’s age (43 years), it is obvious that dentinogenesis is in a less active synthetic state than during the first stages of dentinogenesis. The disjoined cells described in the present study might be the result of a disruption of the intercellular connections reported in transmission electron microscopy10 and SEM,S which means that the odontoblasts might be in a resting state.17 The surface conditions of the circumpulpal dentin have been studied at the coronal level only, during the active stage of dentinogenesis.18 The three-dimensional variations in the structure of the dentinal surface at the coronal and radicular levels have never been compared before. The present data show the dentinal surface to be evenly globular, whereas this
478
Marion et al.
ORAL
7. 8. 9. 10. 11. 12. 13.
REFERENCES 1. Goldman LB, Goldman M, Kronman JH, Lin P.S. Scanning electron microscope study of a new method in endodontic treatment. ORAL SURG ORAL MED ORAL PATHOL 1979;48: 79-83. 2. Cunningham WT, Martin H. A scanning electron microscope evaluation of root canal debridement with the endosonic ultrasonic synergetic system. ORAL SURG ORAL MED ORAL PATHOL 1982;53:527-31.
3. Auther A, Laurichesse JM, Launay Y, Calas P. Etude au microscope a balayage des surfaces radiculaires traittes par les instruments ultrasonores. Rev Fr Endod 1984;3:29-39. 4. Dietschi JM, Ciucchi B, Cergneux M, Holz J, Baume LJ. Preparation du canal radiculaire a I’aide d’ultrasons: etude au MEB. Rev Fr Endod 1984;3:15-27. 5. Jean A, Kerebel B, Kerebel LM. Scanning electron microscope study of the predentin-pulpal border zone in human dentin. SURG ORAL
MED ORAL
MED ORAL
PATHOL
October 1991
type of mineralization has been reported to occur only occasionally along the predentinal forming front.9 The main variations involve the size and shape of the calcospherites, and the number and size of the tubule openings. On the basis of the precise data provided by the present study on the structure of the odontoblasts and of the dentinal surface at the different endodontic levels, it should soon be possible to corroborate the morphologic variations with the cytophysiologic data provided by other techniques and thus acquire a better knowledge of the endodontic functional unit.
ORAL
SURG ORAL
PATHOL
1986;61:392-8.
6. Zerosi C. Compandio di odontoiatrica conservativa terapia,
14. 15. 16. 17. 18.
Ricostruttiva e Endodontia. Milan, Italy: Scienza e Tecnica dentistica, Edizioni Internazionali. 1982:8-26. Seltzer S. Bender IB. The dental .oul~.. 3rd ed. Philadeluhia: JB Lippincott, 1983. Mjor IA, Fejerskov 0. Histology of the human tooth. 3rd ed. Copenhagen, Denmark: 1985, Munskgaard. Bhaskar SN. Orban’s oral histology and embryology. 10th ed. St Louis: CV Mosby, 1986. Frank RM. Etudeau microscopeelectroniquede I’odontoblaste et du canalicule dentinaire humain. Arch Oral Biol 1966;ll: 179-99. Takuma S, Nagai N. Ultrastructure of rat odontoblasts in various stages of their development and maturation. Arch Oral Biol 1971;16:993-1011. Holland CR. Membrane junctions on cat odontoblasts. Arch Oral Biol 1975;20:55 1-2. Viti M, Masi PL. Ultrastructural aspects of interodontoblastic junctions. Bull Group Int Rech Sci Stomatol Odontol 1981;24:61-9. Sasaki T, Nakagawa K, Higashi S. Ultrastructure of odontoblasts in kitten tooth germs as revealed by freeze-fracture. Arch Oral Biol 1982;27:897-904. Scott JH, Symons NBB. Introduction to dental anatomy. 9th ed. Edinburgh: Churchill Livingstone, 1982:216-48. Ten Cate AR. Oral histology: development, structure and function. 2nd ed. St Louis: CV Mosby, 1985. Provenza DV, Seibel W. Oral histology. 2nd ed. Philadelphia: Lea & Febiger, 1982. Lester RS, Boyde A. Electron microscopy of predentinal surfaces. Calcif Tiss Res 1967; 1:44-54.
Reprint
requests:
Dominique Marion Faculte de Chirurgie Dentaire Pl. A. Ricordeau 44042 Nantes Cedex, France
