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Vascular Response of Current and Potential Dental Etchants
JOURNAL OF ENDODONTICS Copyright © 2002 by The American Association of Endodontists
Printed in U.S.A. VOL. 28, NO. 2, FEBRUARY 2002
Vascular Response of Current and Potential Dental Etchants Zafer C. Çehreli, DDS, PhD, Mehmet Ali Onur, PhD, Fügen Tas¸man, DDS, PhD, and Ays¸e Gümrükçüolu, BSc
fibrils at the dentinal tubule wall. Resin penetration into the conditioned dentin results in the formation of intratubular resin tags and a hybrid layer (3). Enhanced adhesion of resin to dentin by etching the dentin with phosphoric acid was first observed in Japan, where dentinal etching has become a fairly common practice (4). In the USA and Europe, however, etching of dentin was discouraged until the 1990s (5). Selective etching of enamel and complete dryness of the tooth substrates were regarded as necessary for optimal bonding and protection of the pulpodentinal complex. Today, however, “all-etch” procedures have been adopted, which involve simultaneous etching of enamel and dentin (4). The effect of etching agents on the pulpodentinal complex is limited by the buffering effect of hydroxyapatite and other dentin components, including collagen, which may act as a barrier that reduces the rate of demineralization (6). However, the placement of acid directly on the mechanically exposed pulp for the purpose of applying a dentin bonding agent to act as a pulp-capping material needs further investigation, because clinical parameters, such as control of pulpal bleeding from the exposure site, may considerably effect pulpal healing. In direct pulp-capping procedures, hemostasis is an important step to biological success, and a clot should not be allowed to form after the cessation of bleeding from the exposure because it impedes pulp healing (7). The clot does not allow the capping material to contact the pulp tissue directly, and subsequently, it will not contact necrosis-producing, degradation products in the clot (7). Although there is recent in vitro evidence that contemporary dentin bonding agents may act as potential candidates for hemorrhage control in direct pulp-capping procedures (8), a survey of the literature fails to demonstrate the hemostatic effect of phosphoric acid on the exposed pulp tissue. The purpose of this study was, therefore, to evaluate the vascular responses produced by dental etchants on the rat carotid artery model.
This study investigated the hemostatic properties of current and potential dentin conditioners by observing the contraction of blood vessels using the rat carotid artery model. Four different agents (3M Scotchbond etchant, NRC, RC-Prep, File-Eze) were used. Dose-dependent contractions/relaxations of the test materials were first compared with epinephrine, followed by administration of papaverine, which was used to reverse epinephrineinduced contractions. In all sequences, the contraction or relaxation forces produced by the test and control materials were recorded using a force displacement transducer. RC-Prep was the only agent to produce epinephrine-like contractions (vasoconstriction) in the rat carotid artery. No contraction could be achieved with 3M etchant. Dosedependent relaxations were observed with 3M etchant, which eventually led to collapse of the vessel wall; afterward, no response could be achieved with administration of adrenalin. File-Eze and NRC also did not cause vasoconstriction. Both materials caused dose-dependent relaxations in the smooth muscle. However, subsequent administration of adrenalin and papaverine caused dosedependent contractions and relaxations, respectively, showing that these etchant effects did not lead to collapse of the vessel wall, as did 3M etchant.
When placing bonded restorations, it is essential to establish a strong and durable bond between the restorative material and the cavity wall to prevent marginal gap formation, postoperative sensitivity, bacterial microleakage, and marginal discoloration (1). The mechanism of current dentin bonding systems is based on resin infiltration into dentin after modification or removal of the smear layer (2). Acid etching of dentin removes the smear layer from the dentinal tubules and from the intertubular dentin, which in addition, is demineralized to a depth of up to 7.5 m (2). The acid also penetrates the opened dentinal tubules, exposing collagen
MATERIALS AND METHODS The conditioning agents used in the study are shown in Table 1. Locally bred female albino rats (Rattus rattus) weighing 300 to 340 g were anesthetized with a mixture of Rompun (2% solution, Bayer, Dormagen, Germany) and Ketalar (Parke Davis, Morris Plains, NJ) (30 mg/kg). The ratio of Rompun and Ketalar in the mixture was 3:1. The left carotid arteries were quickly isolated, cleaned of their connective tissue, and cut into transverse rings of 72
Vol. 28, No. 2, February 2002
Vascular Response of Dental Etchants
73
TABLE 1. Test materials Material
Composition
Manufacturer
3M Scotchbond 37% phosphoric acid etchant NRC
RC-Prep
File-Eze
3M Dental Products Division, St. Paul, MN Maleic acid, itatonic acid, Dentsply DeTrey, water Konstanz, Germany 10% urea peroxide and Medical Products 15% EDTA in a waterLaboratories, soluble vehicle Philadelphia, PA 19% EDTA in an aqueous, Ultradent Products water soluble solution, Inc., South with lubricating base Jordan, UT
3 to 4 mm in length with special care taken not to damage the luminal surface. The rings (n ⫽ 12 per group) were fixed vertically between stainless steel wire hooks and suspended under 0.5 g of resting tension in a tissue bath containing Krebs-bicarbonate solution (pH 7.4) at 37°C, which was continuously aerated with 95% O2 ⫹ CO2. Composition of the Krebs-bicarbonate solution was as follows: NaCl, 118.1 mM; KCl, 4.7 mM; CaCl2, 1.3 mM; NaHCO3, 25 mM; MgCl26H2O 0.5 mM; NaHPO42H2O, 0.9 mM; and glucose. 11 mM. (All chemicals were obtained from Sigma Inc., St. Louis, MO.) Tissues were equilibrated for 30 min. Three different doses (5, 10, and 20 L) of each conditioner were applied and isometric muscle contractions were recorded using a force displacement transducer to which the upper wire was connected. Between each measurement interval, the bathing medium was replaced with 50 ml of Krebs solution. By the end of measurements, three different doses (2, 4, 6 L) of epinephrine (Sigma) were administered in the same manner as to controls and the isometric force measurements were recorded. This was followed by administration of papaverine in three different doses (10, 20, 40 L), and isometric force measurements of the relaxations produced were recorded. In each group, recordings were converted to contraction force (in milligrams). Within each bonding agent group, a Friedman Test with significance set at p ⬍ 0.001, and Wilcoxon signed ranks tests at p ⬍ 0.05 were performed to compare the differences in contraction force for each test dose. The same statistical analyses were also carried out to compare the differences between the contraction force of test materials and that of epinephrine and papaverine. Differences in contraction force between the conditioner groups were determined by Mann-Whitney U Test and with Wilcoxon signed ranks tests. Significance was set at p ⬍ 0.001 and p ⬍ 0.05, respectively. RESULTS The contraction and relaxation forces obtained for the test materials are presented in Figs. 1 and 2 as means and standard deviations. 3M Scotchbond etchant (3M Dental Products Division, St. Paul, MN) produced dose-dependent relaxations in the first two experimental sequences (5 l and 10 l groups). However, no readings could be observed in the 20-l sequence, in which application of the material resulted in a rapid collapse of the vessel wall. This finding was confirmed when subsequent administration of adrenalin in three different doses could elicit no response in the preparation. Therefore, the 3M etchant group was excluded from statistical analyses.
FIG 1. Dose-dependent changes in relaxation force in the carotid artery followed by application of NRC and File-Eze, compared with subsequent administration of papaverine. Bars represent standard deviations.
FIG 2. Dose-dependent contraction forces produced by RC-Prep, compared with subsequently applied epinephrine. Bars represent standard deviations.
NRC (Dentsply DeTrey, Konstanz, Germany) and File-Eze (Ultradent Products Inc., South Jordan, UT) also produced dosedependent relaxations of the rat carotid artery. Unlike the 3M etchant, however, these materials did not cause the vessel to collapse, as confirmed because readings were obtained with subsequent administration of adrenalin and papaverine, which induced contractions and relaxations, respectively. Intramaterial comparison of the differences in relaxation force between the minimal and maximal doses applied for these two materials were found to be statistically significant (p ⬍ 0.01). The relaxation effect of FileEze was greater than that of NRC (Fig. 1). RC-Prep (Medical Products Laboratories, Philadelphia, PA) produced dose-dependent contractions of the carotid artery (p ⬍ 0.01) (Fig. 2). Its effect on the arteries was similar to that of epinephrine (p ⬍ 0.05). Readings that were obtained after the administration of adrenalin and papaverine in the RC-Prep group confirmed that this material caused no collapse in the muscle and that its effect was reversible, as with File-Eze and NRC. DISCUSSION Numerous investigations on enamel and dentin bonding with etchants have focused on comparing different concentrations, viscosities, and etch times of conventional phosphoric acid and weaker acids to improve the bond strength of adhesives to tooth
74
Çehreli et al.
structure. Among these conditioners, a commercially available EDTA gel has been recently shown to produce similar etching patterns to 37% phosphoric acid on cut dentin and enamel (9). Moreover, pretreatment with EDTA not only removes the smear layer, but depletes calcium and phosphate from the surface to open up microporosities much smaller than the dentinal tubules, allowing penetration of hydrophilic resins (10). For these reasons, FileEze and RC-Prep were included in the present study, because both agents contain EDTA. On the other side of the spectrum of etching and chelating agents, the use of self-etching primers, such as NRC, that do not require rinsing and serve simultaneously as conditioner and primer is a recent approach to the simplification of bonding techniques (11). Structural changes in enamel and dentin have been analyzed by scanning electron microscopy after the application of NRC, showing that the demineralization pattern of both enamel and dentin clearly resembles the effects created by phosphoric acid etching (11). NRC is composed of two organic acids dissolved in water (Table 1). Maleic acid acts as a conditioning agent, and itaconic acid behaves as a priming agent with the ability of copolymerizing with the subsequently applied bonding agent. Controversial reports concerning the effects of the total etch technique on exposed pulp tissue prompted our study. Cox et al. (12) and Pameijer and Stanley (13) have indicated that in direct pulp-capping procedures where a total etch technique is used, clinical success strongly depends on complete hemorrhage control before placement of an adhesive system. In this regard, the inherent hemostatic properties of dental etchants would be extremely critical, both biologically and clinically. In their animal studies, Cox et al. (12) used 2.5% sodium hypochlorite (NaOCl) to provide hemorrhage control; Pameijer and Stanley (13) used a cavity disinfectant, Consepsis (Ultradent), for the same purpose. However, the latter authors reported that although good to excellent hemostasis was accomplished with the use of Consepsis after exposure, subsequent application of phosphoric acid caused pulpal bleeding that was difficult to control with the second application of Consepsis. The findings obtained with the 3M etchant in the present study seem to corroborate their observations. When the 3M etchant was administered, relaxation of the arteries was followed by a rapid collapse of the vessel wall, which indicated a chemical denaturation or necrosis, after which no response could be achieved despite the subsequent administration of adrenalin and papaverine. Within the limitations of this study, a clinical implication of this finding may be that more research is indicated at this time before treatment of mechanically exposed vital pulp with phosphoric acid can be considered safe in a total etch procedure; phosphoric acid is not capable of causing vasoconstriction, and the necrosis produced by this acid in the blood vessels may compromise further attempts to control pulpal hemorrhage, either physically or chemically. The relaxation effects that were obtained with NRC and FileEze in this study demonstrated that these conditioners were also not suitable for the control of pulpal hemorrhage. Nevertheless, the vasodilation caused by these etchants was reversible; both conditioners produced no irreversible collapse of the smooth muscle tissue, as it did with 3M etchant. If these materials are to be administered on exposed vital pulp in a total etch procedure, subsequent hemostasis (i.e. with 2.5% NaOCl) would be recommended clinically. The rat carotid artery was used in this study to mimic the smooth muscle contraction in dental pulp vessels, because the behavior of pulpal vessels in response to various known vasoactive agents is not significantly different from this tissue (14). Epinephrine was
Journal of Endodontics
applied as control, because it is known to combine with ␣-adrenergic receptors on arteriolar smooth muscle and cause vasoconstriction (15). The rationale behind administration of papaverine in the present study was to evaluate the reversibility of the contractions obtained with epinephrine. Papaverine is an opium alkaloid, which relaxes many smooth muscles of the body, especially those of blood vessels (16). This relaxation occurs particularly if spasm exists (16). In the present study, RC-Prep was the only conditioner to produce vasoconstriction of the carotid artery, showing that its effect was similar to that of epinephrine and that the material produced no necrotic effect on the smooth muscle tissue, as confirmed by papaverine-induced relaxations. Because both RC-Prep and File-Eze contain EDTA, the vasoconstriction produced by RC-Prep may be attributed to its other constituent, 10% urea peroxide. Most in vivo studies of dentin adhesive systems applied on mechanically exposed pulp tissue of nonhuman primate teeth have reported that resin materials are not toxic and stressed that pulpcapping procedures with dentin adhesives that were performed after pulp etching with acidic agents are a safe therapy, allowing dentin bridging and pulp repair (12). Several authors have reported a strong correlation between microleakage and pulpal reaction (17). However, it is highly unlikely that within 60 or 90 days posttreatment much leakage would occur if contemporary bonding agents were applied to the cavities after a total etch technique (13). Although properly sealed restorations are desirable from the viewpoint of pulpal healing, it is important to distinguish between pulp-cap failure and failure of the restoration subsequently placed over the pulp-capping agent that leads to recurrent pulpitis (18). If a covering restoration is so degraded that a calcium hydroxide base has dissolved, even if not capping a pulp exposure, the amount of microleakage would penetrate even normal patent dentin tubules to create severe pulp pathosis (18). In a recent study (19), pulp inflammation with the absence of bacteria, as well as the reverse, i.e. large amounts of bacteria but no pulp inflammation, was observed. Those authors were unable to attribute cases of pulp necrosis only to the presence of bacteria at the tooth-restoration interface. Therefore, in categorizing the success and failure in pulp-capping procedures, many more factors, including proper hemorrhage control, need to be considered besides the presence or absence of microorganisms. Dr. Çehreli is affiliated with the Department of Pedodontics, Faculty of Dentistry, Hacettepe University, Ankara, Turkey. Drs. Onur and Gümrükçüolu are affiliated with the Department of Biology, Faculty of Science, Hacettepe University, Ankara, Turkey. Dr. Tas¸man is associate professor with the Department of Endodontics, Faculty of Dentistry, Hacettepe University, Ankara, Turkey. Dr. Fügen Tas¸man, Dept. of Endodontics, Faculty of Dentistry, University of Hacettepe, 06100 Sihhiye, Ankara, Turkey.
References 1. Sano H, Takatsu T, Ciucchi B, Horner JA, Pashley DH. Nanoleakage: leakage within the hybrid layer. Oper Dent 1995;20:18 –25. 2. Van Meerbeck B, Inokoshi S, Lambrechts P, Vanherle G. Morphologic aspects of the resin-dentin interdiffusion zone with different dentin adhesive systems. J Dent Res 1992;71:1530 – 40. 3. Nakabayashi N, Kajima K, Masuhara E. The promoter of adhesion by the infiltration of monomers into tooth substances. J Biomed Mater Res 1982;16:265–73. 4. Perdigao J, Lopes M. Dentin bonding: state of the art 1999. Compend Contin Educ Dent 1999;20:1151– 62. 5. Leinfelder KF. Acid etching of dentin: too early to recommend. Quintessence Int 1992;23:229. 6. Uno S, Finger WJ. Effects of acidic conditioners on dentine demineralization and dimension of hybrid layers. J Dent 1996;24:211– 6.
Vol. 28, No. 2, February 2002 7. Kopel HM. Considerations for the direct pulp capping procedure in primary teeth: a review of the literature. ASDC J Dent Child 1992;2:141–9. 8. Onur MA, Tasman F, Cehreli Z, Gumrukcuoglu A. Effect of a fifthgeneration bonding agent on vascular responses in rat carotid artery. J Endodon (in press). 9. Blomlöf JPS, Blomlöf LB, Cederlund AL, Hultenby KR, Lindskog SF. A new concept for etching in restorative dentistry? Int J Periodontics Restorative Dent 1999;19:30 –5. 10. Pashley DH. Dentin bonding: overview of the substrate with respect to adhesive material. J Esthet Dent 1991;3:46 –50. 11. Cehreli ZC, Altay N. Etching effect of 17% EDTA and a non-rinse conditioner (NRC) on primary enamel and dentin. Am J Dent 2000;13:64 – 8. 12. Cox CF, Hafez AA, Akimoto N, Otsuki M, Suzuki S, Tarim B. Biocompatibility of primer, adhesive, and resin composite systems on non-exposed and exposed pulps of non-human primate teeth. Am J Dent 1998;10:55– 63.
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13. Pameijer CH, Stanley HR. The disastrous effects of the “Total Etch” technique in vital pulp capping in primates. Am J Dent 1998;10:45–54. 14. Kim S, Dörscher-Kim J. Hemodynamic regulation of the dental pulp in a low compliance environment. J Endodon 1989;15:404 – 8. 15. Vander AJ, Sherman JH, Luciano DS, eds. Human physiology: the mechanisms of body function. 5th ed. New York: McGraw-Hill, 1990:388 –9. 16. Goodman L, Gilman A, eds. The pharmacological basis of therapeutics. New York: MacMillan, 1941:194 –209. 17. Costa CAS, Mesas AN, Hebling J. Pulp response to direct capping with an adhesive system. Am J Dent 2000;13:81–7. 18. Stanley HR. Criteria for standardizing and increasing credibility of direct pulp capping studies. Am J Dent 1998;11:17–34. 19. Hebling J, Giro EMA, Costa CAS. Human pulp response after an adhesive system application in deep cavities. J Dent 1999;27:557– 64.
Printed in U.S.A. VOL. 28, NO. 2, FEBRUARY 2002
Vascular Response of Current and Potential Dental Etchants Zafer C. Çehreli, DDS, PhD, Mehmet Ali Onur, PhD, Fügen Tas¸man, DDS, PhD, and Ays¸e Gümrükçüolu, BSc
fibrils at the dentinal tubule wall. Resin penetration into the conditioned dentin results in the formation of intratubular resin tags and a hybrid layer (3). Enhanced adhesion of resin to dentin by etching the dentin with phosphoric acid was first observed in Japan, where dentinal etching has become a fairly common practice (4). In the USA and Europe, however, etching of dentin was discouraged until the 1990s (5). Selective etching of enamel and complete dryness of the tooth substrates were regarded as necessary for optimal bonding and protection of the pulpodentinal complex. Today, however, “all-etch” procedures have been adopted, which involve simultaneous etching of enamel and dentin (4). The effect of etching agents on the pulpodentinal complex is limited by the buffering effect of hydroxyapatite and other dentin components, including collagen, which may act as a barrier that reduces the rate of demineralization (6). However, the placement of acid directly on the mechanically exposed pulp for the purpose of applying a dentin bonding agent to act as a pulp-capping material needs further investigation, because clinical parameters, such as control of pulpal bleeding from the exposure site, may considerably effect pulpal healing. In direct pulp-capping procedures, hemostasis is an important step to biological success, and a clot should not be allowed to form after the cessation of bleeding from the exposure because it impedes pulp healing (7). The clot does not allow the capping material to contact the pulp tissue directly, and subsequently, it will not contact necrosis-producing, degradation products in the clot (7). Although there is recent in vitro evidence that contemporary dentin bonding agents may act as potential candidates for hemorrhage control in direct pulp-capping procedures (8), a survey of the literature fails to demonstrate the hemostatic effect of phosphoric acid on the exposed pulp tissue. The purpose of this study was, therefore, to evaluate the vascular responses produced by dental etchants on the rat carotid artery model.
This study investigated the hemostatic properties of current and potential dentin conditioners by observing the contraction of blood vessels using the rat carotid artery model. Four different agents (3M Scotchbond etchant, NRC, RC-Prep, File-Eze) were used. Dose-dependent contractions/relaxations of the test materials were first compared with epinephrine, followed by administration of papaverine, which was used to reverse epinephrineinduced contractions. In all sequences, the contraction or relaxation forces produced by the test and control materials were recorded using a force displacement transducer. RC-Prep was the only agent to produce epinephrine-like contractions (vasoconstriction) in the rat carotid artery. No contraction could be achieved with 3M etchant. Dosedependent relaxations were observed with 3M etchant, which eventually led to collapse of the vessel wall; afterward, no response could be achieved with administration of adrenalin. File-Eze and NRC also did not cause vasoconstriction. Both materials caused dose-dependent relaxations in the smooth muscle. However, subsequent administration of adrenalin and papaverine caused dosedependent contractions and relaxations, respectively, showing that these etchant effects did not lead to collapse of the vessel wall, as did 3M etchant.
When placing bonded restorations, it is essential to establish a strong and durable bond between the restorative material and the cavity wall to prevent marginal gap formation, postoperative sensitivity, bacterial microleakage, and marginal discoloration (1). The mechanism of current dentin bonding systems is based on resin infiltration into dentin after modification or removal of the smear layer (2). Acid etching of dentin removes the smear layer from the dentinal tubules and from the intertubular dentin, which in addition, is demineralized to a depth of up to 7.5 m (2). The acid also penetrates the opened dentinal tubules, exposing collagen
MATERIALS AND METHODS The conditioning agents used in the study are shown in Table 1. Locally bred female albino rats (Rattus rattus) weighing 300 to 340 g were anesthetized with a mixture of Rompun (2% solution, Bayer, Dormagen, Germany) and Ketalar (Parke Davis, Morris Plains, NJ) (30 mg/kg). The ratio of Rompun and Ketalar in the mixture was 3:1. The left carotid arteries were quickly isolated, cleaned of their connective tissue, and cut into transverse rings of 72
Vol. 28, No. 2, February 2002
Vascular Response of Dental Etchants
73
TABLE 1. Test materials Material
Composition
Manufacturer
3M Scotchbond 37% phosphoric acid etchant NRC
RC-Prep
File-Eze
3M Dental Products Division, St. Paul, MN Maleic acid, itatonic acid, Dentsply DeTrey, water Konstanz, Germany 10% urea peroxide and Medical Products 15% EDTA in a waterLaboratories, soluble vehicle Philadelphia, PA 19% EDTA in an aqueous, Ultradent Products water soluble solution, Inc., South with lubricating base Jordan, UT
3 to 4 mm in length with special care taken not to damage the luminal surface. The rings (n ⫽ 12 per group) were fixed vertically between stainless steel wire hooks and suspended under 0.5 g of resting tension in a tissue bath containing Krebs-bicarbonate solution (pH 7.4) at 37°C, which was continuously aerated with 95% O2 ⫹ CO2. Composition of the Krebs-bicarbonate solution was as follows: NaCl, 118.1 mM; KCl, 4.7 mM; CaCl2, 1.3 mM; NaHCO3, 25 mM; MgCl26H2O 0.5 mM; NaHPO42H2O, 0.9 mM; and glucose. 11 mM. (All chemicals were obtained from Sigma Inc., St. Louis, MO.) Tissues were equilibrated for 30 min. Three different doses (5, 10, and 20 L) of each conditioner were applied and isometric muscle contractions were recorded using a force displacement transducer to which the upper wire was connected. Between each measurement interval, the bathing medium was replaced with 50 ml of Krebs solution. By the end of measurements, three different doses (2, 4, 6 L) of epinephrine (Sigma) were administered in the same manner as to controls and the isometric force measurements were recorded. This was followed by administration of papaverine in three different doses (10, 20, 40 L), and isometric force measurements of the relaxations produced were recorded. In each group, recordings were converted to contraction force (in milligrams). Within each bonding agent group, a Friedman Test with significance set at p ⬍ 0.001, and Wilcoxon signed ranks tests at p ⬍ 0.05 were performed to compare the differences in contraction force for each test dose. The same statistical analyses were also carried out to compare the differences between the contraction force of test materials and that of epinephrine and papaverine. Differences in contraction force between the conditioner groups were determined by Mann-Whitney U Test and with Wilcoxon signed ranks tests. Significance was set at p ⬍ 0.001 and p ⬍ 0.05, respectively. RESULTS The contraction and relaxation forces obtained for the test materials are presented in Figs. 1 and 2 as means and standard deviations. 3M Scotchbond etchant (3M Dental Products Division, St. Paul, MN) produced dose-dependent relaxations in the first two experimental sequences (5 l and 10 l groups). However, no readings could be observed in the 20-l sequence, in which application of the material resulted in a rapid collapse of the vessel wall. This finding was confirmed when subsequent administration of adrenalin in three different doses could elicit no response in the preparation. Therefore, the 3M etchant group was excluded from statistical analyses.
FIG 1. Dose-dependent changes in relaxation force in the carotid artery followed by application of NRC and File-Eze, compared with subsequent administration of papaverine. Bars represent standard deviations.
FIG 2. Dose-dependent contraction forces produced by RC-Prep, compared with subsequently applied epinephrine. Bars represent standard deviations.
NRC (Dentsply DeTrey, Konstanz, Germany) and File-Eze (Ultradent Products Inc., South Jordan, UT) also produced dosedependent relaxations of the rat carotid artery. Unlike the 3M etchant, however, these materials did not cause the vessel to collapse, as confirmed because readings were obtained with subsequent administration of adrenalin and papaverine, which induced contractions and relaxations, respectively. Intramaterial comparison of the differences in relaxation force between the minimal and maximal doses applied for these two materials were found to be statistically significant (p ⬍ 0.01). The relaxation effect of FileEze was greater than that of NRC (Fig. 1). RC-Prep (Medical Products Laboratories, Philadelphia, PA) produced dose-dependent contractions of the carotid artery (p ⬍ 0.01) (Fig. 2). Its effect on the arteries was similar to that of epinephrine (p ⬍ 0.05). Readings that were obtained after the administration of adrenalin and papaverine in the RC-Prep group confirmed that this material caused no collapse in the muscle and that its effect was reversible, as with File-Eze and NRC. DISCUSSION Numerous investigations on enamel and dentin bonding with etchants have focused on comparing different concentrations, viscosities, and etch times of conventional phosphoric acid and weaker acids to improve the bond strength of adhesives to tooth
74
Çehreli et al.
structure. Among these conditioners, a commercially available EDTA gel has been recently shown to produce similar etching patterns to 37% phosphoric acid on cut dentin and enamel (9). Moreover, pretreatment with EDTA not only removes the smear layer, but depletes calcium and phosphate from the surface to open up microporosities much smaller than the dentinal tubules, allowing penetration of hydrophilic resins (10). For these reasons, FileEze and RC-Prep were included in the present study, because both agents contain EDTA. On the other side of the spectrum of etching and chelating agents, the use of self-etching primers, such as NRC, that do not require rinsing and serve simultaneously as conditioner and primer is a recent approach to the simplification of bonding techniques (11). Structural changes in enamel and dentin have been analyzed by scanning electron microscopy after the application of NRC, showing that the demineralization pattern of both enamel and dentin clearly resembles the effects created by phosphoric acid etching (11). NRC is composed of two organic acids dissolved in water (Table 1). Maleic acid acts as a conditioning agent, and itaconic acid behaves as a priming agent with the ability of copolymerizing with the subsequently applied bonding agent. Controversial reports concerning the effects of the total etch technique on exposed pulp tissue prompted our study. Cox et al. (12) and Pameijer and Stanley (13) have indicated that in direct pulp-capping procedures where a total etch technique is used, clinical success strongly depends on complete hemorrhage control before placement of an adhesive system. In this regard, the inherent hemostatic properties of dental etchants would be extremely critical, both biologically and clinically. In their animal studies, Cox et al. (12) used 2.5% sodium hypochlorite (NaOCl) to provide hemorrhage control; Pameijer and Stanley (13) used a cavity disinfectant, Consepsis (Ultradent), for the same purpose. However, the latter authors reported that although good to excellent hemostasis was accomplished with the use of Consepsis after exposure, subsequent application of phosphoric acid caused pulpal bleeding that was difficult to control with the second application of Consepsis. The findings obtained with the 3M etchant in the present study seem to corroborate their observations. When the 3M etchant was administered, relaxation of the arteries was followed by a rapid collapse of the vessel wall, which indicated a chemical denaturation or necrosis, after which no response could be achieved despite the subsequent administration of adrenalin and papaverine. Within the limitations of this study, a clinical implication of this finding may be that more research is indicated at this time before treatment of mechanically exposed vital pulp with phosphoric acid can be considered safe in a total etch procedure; phosphoric acid is not capable of causing vasoconstriction, and the necrosis produced by this acid in the blood vessels may compromise further attempts to control pulpal hemorrhage, either physically or chemically. The relaxation effects that were obtained with NRC and FileEze in this study demonstrated that these conditioners were also not suitable for the control of pulpal hemorrhage. Nevertheless, the vasodilation caused by these etchants was reversible; both conditioners produced no irreversible collapse of the smooth muscle tissue, as it did with 3M etchant. If these materials are to be administered on exposed vital pulp in a total etch procedure, subsequent hemostasis (i.e. with 2.5% NaOCl) would be recommended clinically. The rat carotid artery was used in this study to mimic the smooth muscle contraction in dental pulp vessels, because the behavior of pulpal vessels in response to various known vasoactive agents is not significantly different from this tissue (14). Epinephrine was
Journal of Endodontics
applied as control, because it is known to combine with ␣-adrenergic receptors on arteriolar smooth muscle and cause vasoconstriction (15). The rationale behind administration of papaverine in the present study was to evaluate the reversibility of the contractions obtained with epinephrine. Papaverine is an opium alkaloid, which relaxes many smooth muscles of the body, especially those of blood vessels (16). This relaxation occurs particularly if spasm exists (16). In the present study, RC-Prep was the only conditioner to produce vasoconstriction of the carotid artery, showing that its effect was similar to that of epinephrine and that the material produced no necrotic effect on the smooth muscle tissue, as confirmed by papaverine-induced relaxations. Because both RC-Prep and File-Eze contain EDTA, the vasoconstriction produced by RC-Prep may be attributed to its other constituent, 10% urea peroxide. Most in vivo studies of dentin adhesive systems applied on mechanically exposed pulp tissue of nonhuman primate teeth have reported that resin materials are not toxic and stressed that pulpcapping procedures with dentin adhesives that were performed after pulp etching with acidic agents are a safe therapy, allowing dentin bridging and pulp repair (12). Several authors have reported a strong correlation between microleakage and pulpal reaction (17). However, it is highly unlikely that within 60 or 90 days posttreatment much leakage would occur if contemporary bonding agents were applied to the cavities after a total etch technique (13). Although properly sealed restorations are desirable from the viewpoint of pulpal healing, it is important to distinguish between pulp-cap failure and failure of the restoration subsequently placed over the pulp-capping agent that leads to recurrent pulpitis (18). If a covering restoration is so degraded that a calcium hydroxide base has dissolved, even if not capping a pulp exposure, the amount of microleakage would penetrate even normal patent dentin tubules to create severe pulp pathosis (18). In a recent study (19), pulp inflammation with the absence of bacteria, as well as the reverse, i.e. large amounts of bacteria but no pulp inflammation, was observed. Those authors were unable to attribute cases of pulp necrosis only to the presence of bacteria at the tooth-restoration interface. Therefore, in categorizing the success and failure in pulp-capping procedures, many more factors, including proper hemorrhage control, need to be considered besides the presence or absence of microorganisms. Dr. Çehreli is affiliated with the Department of Pedodontics, Faculty of Dentistry, Hacettepe University, Ankara, Turkey. Drs. Onur and Gümrükçüolu are affiliated with the Department of Biology, Faculty of Science, Hacettepe University, Ankara, Turkey. Dr. Tas¸man is associate professor with the Department of Endodontics, Faculty of Dentistry, Hacettepe University, Ankara, Turkey. Dr. Fügen Tas¸man, Dept. of Endodontics, Faculty of Dentistry, University of Hacettepe, 06100 Sihhiye, Ankara, Turkey.
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