Reactive changes in dental tissues in response to curing light exposure: An experimental study

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BACKGROUND

Modern dentistry routinely employs light-curable restorative materials. The introduction of light-curing devices has marked a significant advancement in the quality of dental care. As with all medical devices, their safety must be ensured [1]. Light-curing protocols are primarily developed with regard to polymerization efficacy (completeness of cure). However, a notable drawback of curing light exposure is the potential for thermal injury to the dental pulp and surrounding tissues. There is a clear need for experimental validation of differentiated light-curing protocols, specifically intensity and exposure time, to minimize tissue impact [2]. No similar studies were identified in the available databases.

AIM: To assess the effect of curing light on dental tissues.

METHODS

Study Design

An experimental simulation of curing light application in dental restorative procedures was conducted. The study included 50 rats randomized into four groups: Group 1 (control, n=5) and Groups 2–4 (treatment, n=15 each). Animals in the control group were not exposed to curing light. In treatment groups, the mandibular incisors were irradiated twice with a dental curing device emitting light at 395–480 nm. Three light intensity modes were applied: Group 2 was exposed to 1,000 mW/cm², Group 3 to 1,400 mW/cm², and Group 4 to 3,200 mW/cm². The animals were sacrificed by decapitation after 7 days in Group 1 and after 1, 3, and 7 days in Groups 2, 3, and 4, respectively. A total of 100 teeth (two mandibular incisors per animal) were extracted for pathomorphological examination. From these specimens, 320 histological sections (including enamel, dentin, and pulp) were prepared, stained, and analyzed.

Study Setting

The experiment was carried out at Smolensk State Medical University. Animals were housed in a vivarium under standard sanitary conditions in labeled enclosures¹. For procedural sedation and decapitation, animals received intraperitoneal premedication with 2% xylazine hydrochloride solution (Xyla, Interchemie Werken “De Adelaar” B.V., Netherlands) at a dose of 0.15 mL/kg, followed by anesthesia with Zoletil (Virbac Santé Animale, France) at a dose of 0.01 mg/kg [3].

Study Duration

Dental tissue responses to curing light were assessed dynamically 1, 3, and 7 days post-exposure. The experiment was conducted in 2023.

Measurement Protocol

The extracted teeth were decalcified for 18 days in trichloroacetic acid. Following initial fixation, the specimens were trimmed, re-fixed above the liquid level in trichloroacetic acid for 24 hours, and embedded in paraffin. Histological sections 5–8 µm thick were prepared using a microtome. Deparaffinization was performed by treating the sections with three changes of xylene for 5 minutes each. Dehydration was achieved by sequential immersion in four ethanol concentrations (absolute, 96%, 70%, and 56%) for 3 minutes each. The material was fixed in 15% aqueous neutral buffered formalin and paraffin-embedded. Histological staining included hematoxylin and eosin, Van Gieson, Alcian blue, and periodic acid–Schiff (PAS) reaction. The vascular bed and its components were examined using the Gab—Dyban technique. Standardized comparative microscopic analysis of dental tissues (enamel, dentin, and pulp) was performed using the Axio Imager A2 imaging system (Carl Zeiss, Germany) at ×100 and ×200 magnification. Digital images were acquired using the Axiocam 506 color camera for light microscopy (Carl Zeiss, Germany).

Main Study Outcome

The primary objective was to test the hypothesis that curing light exposure, under manufacturer-recommended settings for composite restorative materials, induces reactive changes in dental tissues. The study also sought to characterize the underlying pathomorphological features of these responses.

Additional Study Outcomes

An additional outcome was to assess the reversibility of the observed tissue changes and their resolution.

Subgroup Analysis

The study included 8-month-old male rats. Group 1 served as the control, while Groups 2 through 4 received curing light exposure. Differences between the control and treatment groups were assessed by analyzing intragroup changes following exposure to diode curing light.

Outcomes Registration

Outcomes were assessed using descriptive statistics and semiquantitative evaluation, with ranked scoring of histopathologic features.

Given the inherent subjectivity of microscopic evaluation, determined by the study subject and observer, both intraobserver and interobserver validation methods were employed to minimize bias.

Ethics Approval

The study design was justified by weighing the potential harm to animals against the prospective benefit to human health. The use of animals was limited to the minimum necessary to achieve scientific validity. All procedures adhered to current legal and ethical standards². The study was approved by the Ethics Committee of Smolensk State Medical University (Minutes No. 2 of September 23, 2022).

Statistical Analysis

Descriptive statistics were used to assess the findings.

RESULTS

Primary Results

Histologic examination of dental tissues in the control group (Group 1) revealed preserved structural components: enamel, dentin, and pulp. Enamel, physiologically thin in this species, covered the crown portion of the tooth. Dentin, a connective tissue, consisted of fibroblasts and odontoblasts within an extracellular matrix composed of procollagen, collagen, and reticular fibers. At the boundary with predentin, odontoblasts were arranged in rows of 6–8 cells within the coronal pulp. These cells varied in shape from prismatic or pear-shaped to cuboidal. Beneath the odontoblast layer in the coronal portion of the tooth, the subodontoblastic layer consists of the cell-free (zone of Weil) and cell-rich zones containing fibroblasts, lymphocytes, and poorly differentiated cells capable of differentiating into odontoblasts. The pulp exhibited rich innervation and vascularization. Concentric radial dentin layers followed the odontoblast layer, with the dentin layer approximately 1.5 times thicker than the pulp. Overall, the dental microstructure resembled that of a human tooth [4], with two distinguishing features: thinner enamel and more pronounced pulp vascularity.

In Groups 2–4, teeth were exposed to curing light (395–480 nm). In Group 2 (1,000 mW/cm²), no signs of pulp inflammation were observed on Day 1. By Day 3, individual lymphocytes and polymorphonuclear leukocytes appeared in the loose connective tissue stroma of the pulp. Microvascular blood filling was heterogeneous; some vessels exhibited dilated lumens.

In Group 3 (1,400 mW/cm², 3-second exposure), histologic architecture was preserved on Day 1 (Fig. 1), and findings were largely within normal limits. However, heterogeneous blood filling of the pulp microvasculature was noted. The odontoblast layer appeared thinned and sparsely populated. Capillary walls exhibited plasmorrhagia, with occasional stasis in the lumen; endothelial cells remained intact. On Day 3, mild perivascular lymphocytic infiltration was observed in the pulp stroma (Fig. 2).

 

Fig. 1. Transverse histological section of a tooth (third experimental group, day 1). Hematoxylin and eosin staining; ×100.

 

Fig. 2. Perivascular infiltration in the dental pulp (third experimental group, day 3). Hematoxylin and eosin staining; ×200.

 

In Group 4 (3,200 mW/cm²), Day 1 findings showed preserved overall dental structure with scattered cellular infiltration of the pulp and good vascular filling (Fig. 3). Lymphocytes were noted in the loose connective tissue stroma. On Day 3, dental hard tissues remained structurally intact. Pulp microvasculature exhibited uneven blood filling, and sparse lymphocytes were present in the stroma. PAS staining revealed weak glycogen positivity in the pulp, with higher glycogen content in circumpulpal dentin compared to mantle dentin (Fig. 4). Pulp vessels remained engorged, with areas of vascular stasis.

 

Fig. 3. Transverse histological section of a tooth (fourth experimental group, day 3). Intensive presence of glycogen in the tooth tissues around the pulp. Staining using the PAS reaction; ×100.

 

Fig. 4. Transverse histological section of a tooth (fourth experimental group, day 1). The degree of blood filling of the dental pulp is shown. Staining according to Gabu–Dyban; ×200.

 

Histochemical staining showed preserved structure of hard dental tissues in all groups at all time points. The dentin exhibited a radial structural pattern (see Fig. 1).

Secondary Results

The findings suggest that reactive changes in the dental pulp induced by curing light exposure are reversible.

On Day 7, histologic features of the pulp in Groups 2 and 3 approximated normal. The odontoblast layer maintained its histoarchitectonic integrity, and microvasculature showed vascular engorgement without evidence of inflammatory infiltrate.

In Group 4, Day 7 histology remained consistent with that observed on Days 1 and 3. The edematous stroma contained scattered lymphocytes, plasma cells, and polymorphonuclear leukocytes. PAS staining continued to demonstrate increased glycogen content in the dentin adjacent to the pulp. The pulp vasculature remained markedly engorged.

Adverse Events

No adverse events were observed during the study.

DISCUSSION

The experimental findings indicate a risk of negative photochemical reactions in the pulp following curing light exposure during treatment of patients with hard dental tissue pathologies. Histopathologic changes in the pulp exhibited features of acute inflammation, most pronounced on Day 3, and were interpreted as a protective response to irritation [5, 6]. The most significant alterations involved the microvasculature, including increased and heterogeneous blood engorgement, plasmorrhagia of capillary walls, and stasis within their lumens. Although this condition typically does not require specific treatment, it may cause transient postoperative discomfort in some patients following light curing.

There were no morphological changes in hard dental tissues.

The initial pulp response to curing light exposure in experimental animals was uniform and appeared independent of the technical specifications of the curing devices. By Day 7, reactive changes in the pulp resolved completely in groups exposed to diode light at 1,000 and 1,400 mW/cm². At this point, the pulp morphology in animals exposed to 3,200 mW/cm² light approximated baseline findings, with residual perivascular cellular infiltration and persistent capillary congestion.

CONCLUSION

These data regarding the effects of curing light on the dental pulp have practical implications for selecting curing protocols during composite restorations, aiming to minimize the risk of unexpected tissue reactions.

ADDITIONAL INFORMATION

Funding sources: This work was not supported by any external sources.

Disclosure of interests: The authors declare no explicit or potential conflicts of interests associated with the publication of this article.

Author contributions: V.R. Shashmurina: conceptualization, review, writing; Valery M. Kiselev, investigation; L.N. Grishenkova: study coordination, editing; A.S. Novikov: statistical analysis, data integration and analysis. All authors confirm that their authorship meets the international ICMJE criteria (all authors made substantial contributions to the conceptualization, investigation, and manuscript preparation, and reviewed and approved the final version prior to publication).

 

¹ GOST 33215-2014. Guidelines for accommodation and care of animals. Environment, housing, and management. Available at: https://docs.cntd.ru/document/1200127789

² GOST 33044-2014. International standart. Principles of good laboratory practice. Available at: https://docs.cntd.ru/document/1200115791

作者简介

Victoria Shashmurina

Smolensk State Medical University

编辑信件的主要联系方式.
Email: Shahmurina@yandex.ru
ORCID iD: 0000-0001-5216-7521
SPIN 代码: 4199-4204

MD, Dr. Sci. (Medcine), Professor

俄罗斯联邦, 28 Krupskoy street, 214019 Smolensk

Valery Kiselev

Smolensk State Medical University

Email: kiselevm23@mail.ru
ORCID iD: 0000-0002-8678-8606
俄罗斯联邦, 28 Krupskoy street, 214019 Smolensk

Liudmila Grishenkova

Smolensk State Medical University

Email: lgrish@gmail.com
ORCID iD: 0009-0003-8328-6947
SPIN 代码: 7516-4492

MD, Cand. Sci. (Medicine), Associate Professor

俄罗斯联邦, 28 Krupskoy street, 214019 Smolensk

Alexandr Novikov

Smolensk State Medical University

Email: a9516900315@yandex.ru
ORCID iD: 0009-0008-3868-0562
SPIN 代码: 2039-8246

MD, Cand. Sci. (Medicine), Associate Professor

俄罗斯联邦, 28 Krupskoy street, 214019 Smolensk

参考

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