Pulp and apical tissue response to deep caries in immature teeth: A histologic and histobacteriologic study

Abstract

Descriptions of the pathologic changes in the pulp and associated apical structures of human immature teeth in response to deep caries are lacking in the literature.

Objectives

This article describes the histologic events associated with the radicular pulp and the apical tissues of human immature teeth following pulp inflammation and necrosis.

Methods

Twelve immature teeth with destructive caries lesions were obtained from 8 patients. Two intact immature teeth served as controls. Teeth were extracted for reasons not related to this study and immediately processed for histopathologic and histobacteriologic analyses. Serial sections were examined for the pulp conditions and classified as reversible or irreversible pulp inflammation, or pulp necrosis. Other histologic parameters were also evaluated.

Results

In the 3 cases with reversible pulp inflammation, tissue in the pulp chamber showed mild to moderate inflammation and tertiary dentin formation related to tubules involved in the caries process. Overall, the radicular pulp tissue, apical papilla and Hertwig’s epithelial root sheath (HERS) exhibited characteristics of normality. In the 3 cases with irreversible pulp inflammation, the pulps were exposed and severe inflammation occurred in the pulp chamber, with minor areas of necrosis and infection. Large areas of the canal walls were free from odontoblasts and lined by an atubular mineralized tissue. The apical papilla showed extremely reduced cellularity or lack of cells and HERS was discontinuous or absent. In the 6 cases with pulp necrosis, the coronal and radicular pulp tissue was necrotic and colonized by bacterial biofilms. The apical papilla could not be discerned, except for one case. HERS was absent in the necrotic cases.

Conclusion

While immature teeth with reversible pulpitis showed histologic features almost similar to normal teeth in the canal and in the apical region, those with irreversible pulpitis and necrosis exhibited significant alterations not only in the radicular pulp but also in the apical tissues, including the apical papilla and HERS.

Clinical significance

Alterations in the radicular pulp and apical tissues help explain the outcome of current regenerative/reparative therapies and should be taken into account when devising more predictable therapeutic protocols for teeth with incomplete root formation.

Introduction

Immature teeth are characterized by open apices with thin dentin walls. In early stages of root formation, the proliferating end of the Hertwig’s epithelial root sheath (HERS), the union of inner and external enamel epithelium, bends at a near 45° angle. This structure is referred to as epithelial diaphragm and is believed to be responsible for root dentin formation through signaling to undifferentiated mesenchymal cells present in the dental papilla . As the differentiated odontoblasts lay down the primary dentin, the dental papilla becomes encased within the dentin structure. In advanced stages of root development the volume of dental papilla is considerably reduced, restricted to only the apical portion of the root. This structure has been named apical papilla . The apical papilla harbors stem cells that are believed to be capable of differentiating into primary odontoblasts for formation of root dentin . In histologic sections stained with hematoxylin and eosin, the apical papilla appears as an eosinophilic tissue with large cells, separated from the rest of the pulp by a cell-rich zone . HERS can usually be observed layering the external apical surface, sometimes enveloping the thin apical dentin walls.

While tissue response to caries in fully formed teeth has been well documented , descriptions of the pathologic changes occurring in the pulp and associated immature apical structures, as a consequence of deep caries affecting human young teeth, are lacking in the literature, most likely due to availability of the samples. The purpose of this article is to describe histologic events taking place in the radicular pulp and in the apical portion of human immature roots, including the apical papilla and HERS, following pulp inflammation and necrosis. Knowledge of the pulp and apical tissue response in these conditions is relevant to understand and predict the healing mechanisms following application of current regeneration/reparative therapies and has also the potential to guide the development of strategies to predictably treat human immature teeth.

Materials and methods

Twelve teeth with destructive caries lesions (4 maxillary first molars, 6 mandibular first molar, 1 mandibular second molar, and 1 maxillary second premolar) were obtained from 8 patients (5 males, 3 females; ages ranging from 7 to 15 years; mean age, 9.7 years). Case description and clinical diagnosis are provided in Table 1 . Patients were brought by their parents to the Department of Maxillofacial Surgery at the School of Dentistry. The teeth were extracted because they were deemed nonrestorable, or their parents requested extraction. All teeth exhibited incomplete root formation on the basis of panoramic and periapical radiographs. Two intact immature teeth (one maxillary second premolar extracted for orthodontic reasons, and one mandibular third molar extracted because of pericoronitis) were used as controls. Before tooth extraction, written informed consent was obtained from patient’s parents. The study was approved by the Institutional Review Board of New York University Medical Center.

Table 1
Teeth with immature roots and different clinical and histopathological diagnosis. Case description.
Sample Age Sex Tooth Pulp diagnosis Periapical diagnosis Histological pulp diagnosis Odontoblast changes in canal Tertiary dentin in canals Changes in apical papilla Changes in HERS
1 10 M #14 Pulp necrosis Symptomatic AP Irreversible pulpitis Yes Yes Yes Yes
2 9 M #30 Unknown Symptomatic AP Reversible pulpitis Yes Yes No No
3 10 M #3 Irreversible pulpitis Symptomatic AP Irreversible pulpitis Yes Yes Yes Yes
4 13 F #4 Reversible pulpitis Normal apical tissue Reversible pulpitis Yes Yes Yes No
5 10 M #31 Pulp necrosis Acute apical abscess Pulp necrosis Yes Yes Yes Yes
6 9 F #19 Pulp necrosis Acute apical abscess Pulp necrosis Yes Yes Yes Yes
7 9 M #19 Unknown Symptomatic apical periodontitis Reversible pulpitis Yes Yes Yes No
8 8 M #14 Previously initiated therapy Unknown Pulp necrosis Yes Yes Yes Yes
9 9 M #3 Unknown. Indirect pulp capping Symptomatic apical periodontitis Irreversible pulpitis (pulp polyp present) Yes Yes Yes Yes
10 15 M #31 Pulp necrosis Asymptomatic apical periodontitis Pulp necrosis Yes Yes Yes Yes
11 7 F #19 Pulp necrosis Unknown Pulp necrosis Yes Yes Yes Yes
12 7 F #30 Irreversible pulpitis Normal apical tissue Pulp necrosis Yes Yes Yes Yes
Note : 1: sample ID 1 and 3 were collected from the same patient; 2: sample ID 2, 7, and 8 were collected from the same patient; 3: sample ID 11 and 12 were collected from the same patient.

Immediately after extraction, the teeth were fixed in 10% neutral buffered formalin solution. Demineralization was carried out in an aqueous solution consisting of a mixture of 22.5% (vol/vol) formic acid and 10% (wt/vol) sodium citrate for 4 weeks. The end point was determined radiographically. Following demineralization, maxillary molar teeth (cases #1, 3, 8, and 9; Table 1 ) were separated into two portions with a sharp razor blade on a mesio-distal plane, approximately at the center of the pulp chamber. One portion, encompassing the two buccal root and half crown, was embedded so that longitudinal sections could be cut on a mesio-distal plane. For the second portion, encompassing the palatal root and the other half crown, longitudinal sections were cut on a bucco-lingual plane, in an effort to obtain, in curved roots, sections including the pulp chamber and the entire canal course from orifice to foramen.

Mandibular molars with diverging roots were separated at the furcation on a bucco-lingual direction (cases #2, 5, and 6; Table 1 ). The two portions, encompassing mesial and distal root respectively, were embedded so that longitudinal sections were cut on a mesio-distal plane. The remaining mandibular molars (cases #7, 10, 11, and 12; Table 1 ) were embedded in one piece, and longitudinal sections on a mesio-distal plane were cut.

After demineralization, the specimens were washed in running water for 24 h, dehydrated in ascending grades of ethanol, cleared in xylene, and infiltrated and embedded in paraffin (melting point of 56 °C) according to standard procedures.

Serial sections were taken with the microtome set at 4–5 μm. Several hundred sections were taken for each specimen, until the wide apical canals and foraminal openings were exhausted. Every 5th slide was stained with hematoxylin and eosin (H&E) for screening purposes and assessment of inflammation. A modified Brown and Brenn technique for staining bacteria was used for selected slides. The remaining slides were stained as needed.

Irrespective of the clinical diagnosis, the teeth were histologically classified into three categories according to a slight modification of the criteria described elsewhere ( Table 1 ):

  • a)

    Reversible pulp disease . This group included specimens with evidence of moderate or severe chronic inflammation confined to the coronal pulp. Lymphocytes and plasma cells are seen gathered in varying concentrations beneath the deepest areas of caries penetration. Absence of even minimal areas of coagulation or liquefaction necrosis. Bacteria are seen in large numbers in the carious dentin, but are absent in the pulp tissue.

  • b)

    Irreversible pulp disease . At least one area, even very small, of the coronal pulp tissue has undergone liquefaction or coagulation, surrounded by masses of live and dead polymorphonuclear neutrophils (PMNs). Bacterial aggregations/biofilms are observed colonizing the necrotic pulp tissue or the adjacent dentin walls. Peripherally, concentrations of chronic inflammatory cells (lymphocytes, plasma cells and macrophages) form a dense halo around these central zones of abscess.

  • c)

    Pulp necrosis . The entire pulp tissue, or large part of it, has undergone necrosis with bacterial aggregations/biofilms colonizing the necrotic tissue and the adjacent dentin walls.

In addition, the following aspects were specifically investigated and recorded ( Table 1 ):

  • 1.

    Changes in the regularity of the odontoblast layer on the root canal walls;

  • 2.

    Presence of tertiary dentin formed on the root canal walls;

  • 3.

    Changes in cellularity of the apical papilla;

  • 4.

    Changes in the HERS surrounding the apical contour.

The worst reactions were recorded for each tooth.

Materials and methods

Twelve teeth with destructive caries lesions (4 maxillary first molars, 6 mandibular first molar, 1 mandibular second molar, and 1 maxillary second premolar) were obtained from 8 patients (5 males, 3 females; ages ranging from 7 to 15 years; mean age, 9.7 years). Case description and clinical diagnosis are provided in Table 1 . Patients were brought by their parents to the Department of Maxillofacial Surgery at the School of Dentistry. The teeth were extracted because they were deemed nonrestorable, or their parents requested extraction. All teeth exhibited incomplete root formation on the basis of panoramic and periapical radiographs. Two intact immature teeth (one maxillary second premolar extracted for orthodontic reasons, and one mandibular third molar extracted because of pericoronitis) were used as controls. Before tooth extraction, written informed consent was obtained from patient’s parents. The study was approved by the Institutional Review Board of New York University Medical Center.

Table 1
Teeth with immature roots and different clinical and histopathological diagnosis. Case description.
Sample Age Sex Tooth Pulp diagnosis Periapical diagnosis Histological pulp diagnosis Odontoblast changes in canal Tertiary dentin in canals Changes in apical papilla Changes in HERS
1 10 M #14 Pulp necrosis Symptomatic AP Irreversible pulpitis Yes Yes Yes Yes
2 9 M #30 Unknown Symptomatic AP Reversible pulpitis Yes Yes No No
3 10 M #3 Irreversible pulpitis Symptomatic AP Irreversible pulpitis Yes Yes Yes Yes
4 13 F #4 Reversible pulpitis Normal apical tissue Reversible pulpitis Yes Yes Yes No
5 10 M #31 Pulp necrosis Acute apical abscess Pulp necrosis Yes Yes Yes Yes
6 9 F #19 Pulp necrosis Acute apical abscess Pulp necrosis Yes Yes Yes Yes
7 9 M #19 Unknown Symptomatic apical periodontitis Reversible pulpitis Yes Yes Yes No
8 8 M #14 Previously initiated therapy Unknown Pulp necrosis Yes Yes Yes Yes
9 9 M #3 Unknown. Indirect pulp capping Symptomatic apical periodontitis Irreversible pulpitis (pulp polyp present) Yes Yes Yes Yes
10 15 M #31 Pulp necrosis Asymptomatic apical periodontitis Pulp necrosis Yes Yes Yes Yes
11 7 F #19 Pulp necrosis Unknown Pulp necrosis Yes Yes Yes Yes
12 7 F #30 Irreversible pulpitis Normal apical tissue Pulp necrosis Yes Yes Yes Yes
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Jun 19, 2018 | Posted by in General Dentistry | Comments Off on Pulp and apical tissue response to deep caries in immature teeth: A histologic and histobacteriologic study
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