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Development and validation of an in vitro model for measurements of cervical root dentine permeability.

Jungbluth H, Attin T, Buchalla W - Clin Oral Investig (2014)

Bottom Line: In experiment 2, permeability was different within the specimens and decreased highly linearly with time, allowing the prediction of future values.In experiment 3, Seal&Protect® completely occluded dentinal tubules. elmex fluid® increased tubular permeability by about 30 % compared to control.The described in vitro model allows evaluation of potential agents for the treatment of DH at the clinically relevant cervical region of human teeth.

View Article: PubMed Central - PubMed

Affiliation: Department of Preventive Dentistry, Periodontology, and Cariology, University of Zürich Center for Dental Medicine, Plattenstrasse 11, 8032, Zürich, Switzerland, holger.jungbluth@zzm.uzh.ch.

ABSTRACT

Objectives: The aim of this series of studies was the development and validation of a new model for evaluation of dentinal hypersensitivity (DH) therapies.

Materials and methods: Roots from extracted human teeth were sealed with a flowable composite. In the cervical area, a 3-mm-wide circular window was ground through the seal 1 mm deep into dentine. The pulp lumen was connected to a reservoir of artificial dentinal fluid (ADF) containing protein, mineral salts and methylene blue. At increased pulpal pressure, the ADF released through the said window was collected in containers each with 20 ml of physiologic saline for a consecutive series of 30-min intervals and ADF concentration (absorption) was determined photometrically. The model was verified by three experiments. In experiment 1, the lower limit of quantification (LLoQ, coefficient of variation = 20 % and difference of 5 standard deviations (SD) from blank) of ADF in physiologic saline was determined by measuring the absorption of 15 dilutions of ADF in physiologic saline (containing 0.625 ng to 12.5 μg methylene blue/ml) photometrically for ten times. In experiment 2, long-term linearity of ADF perfusion/outflow was investigated using 11 specimens. The ADF released through the window was collected in the said containers separately for each consecutive interval of 30 min for up to 240 min. Absorption was determined and analysed by linear regression over time. In experiment 3, perfusion before (2×) and after single treatment according to the following three groups was measured: BisGMA-based sealant (Seal&Protect®), an acidic fluoride solution (elmex fluid®) and control (no treatment).

Results: In experiment 1, the LLoQ was 0.005 μg methylene blue/ml. In experiment 2, permeability was different within the specimens and decreased highly linearly with time, allowing the prediction of future values. In experiment 3, Seal&Protect® completely occluded dentinal tubules. elmex fluid® increased tubular permeability by about 30 % compared to control.

Conclusions: A model comprising the use of artificial dentinal fluid was developed and validated allowing screening of therapeutic agents for the treatment of DH through reliable measurement of permeability of cervical root dentine.

Clinical relevance: The described in vitro model allows evaluation of potential agents for the treatment of DH at the clinically relevant cervical region of human teeth.

No MeSH data available.


Related in: MedlinePlus

Cycling of the pressure within the specimen with time. The test begins at high pressure (525 hPa). When a continuous flow of ADF is observed, the specimen is placed into tube 1 and time measurement is started (time point 0 min). In each tube, the specimen rests for 30 min at high pressure, followed by a 15-min low-pressure interval in physiologic saline as an intermediate storage medium in another tube (not shown). End point of experiment 3 is signed by alpha (α) and of experiment 2 by omega (Ω)
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Fig3: Cycling of the pressure within the specimen with time. The test begins at high pressure (525 hPa). When a continuous flow of ADF is observed, the specimen is placed into tube 1 and time measurement is started (time point 0 min). In each tube, the specimen rests for 30 min at high pressure, followed by a 15-min low-pressure interval in physiologic saline as an intermediate storage medium in another tube (not shown). End point of experiment 3 is signed by alpha (α) and of experiment 2 by omega (Ω)

Mentions: In the second experiment, the stability and drift of the perfusion through the dentine was assessed over a longer time period. A pressure of about 525 hPa was established in the pulpal space of 11 single specimens (n = 11) that were seated in a storage vessel filled with physiologic saline by increasing the pressure in the ADF reservoir to 500 hPa. When a continuous stream of ADF could be seen passing through the specimen window, the experiment was started (Figs. 2 and 3). Hitherto, the specimens were rinsed with physiologic saline from outside, gently dried and immediately put into a new centrifuge tube (tube 1) with 20 ml of fresh physiologic saline, and the time was measured beginning with this starting point (time = 0 min in Fig. 3). The specimens were kept in the tube for 30 min and then removed (at time point 30 min in Fig. 3) and stored transiently at reduced, environmental pressure conditions (=25 hPa of the apparatus) in a storage vessel also containing fresh physiologic saline. After 15 min of storage, the specimens were put into a new centrifuge tube (tube 2, at time point 45 min) with 20 ml fresh saline and the pressure was raised again to 525 hPa. After another 30 min (at time point 75 min), the specimens were removed from tube 2. These alternating pressure conditions were repeated five times (up to time point 255 min), and the absorption caused by the dye stained ADF in each centrifuge tube (tubes 1 to 6) was measured with the photometer at a wavelength of 664 nm. During perfusion time, the absence of ADF perfusion beyond the previously described window was determined visually. In case of any leakage beyond this window, e.g. through a gap between specimen holder and root, the respective specimen was discarded.Fig. 2


Development and validation of an in vitro model for measurements of cervical root dentine permeability.

Jungbluth H, Attin T, Buchalla W - Clin Oral Investig (2014)

Cycling of the pressure within the specimen with time. The test begins at high pressure (525 hPa). When a continuous flow of ADF is observed, the specimen is placed into tube 1 and time measurement is started (time point 0 min). In each tube, the specimen rests for 30 min at high pressure, followed by a 15-min low-pressure interval in physiologic saline as an intermediate storage medium in another tube (not shown). End point of experiment 3 is signed by alpha (α) and of experiment 2 by omega (Ω)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4232750&req=5

Fig3: Cycling of the pressure within the specimen with time. The test begins at high pressure (525 hPa). When a continuous flow of ADF is observed, the specimen is placed into tube 1 and time measurement is started (time point 0 min). In each tube, the specimen rests for 30 min at high pressure, followed by a 15-min low-pressure interval in physiologic saline as an intermediate storage medium in another tube (not shown). End point of experiment 3 is signed by alpha (α) and of experiment 2 by omega (Ω)
Mentions: In the second experiment, the stability and drift of the perfusion through the dentine was assessed over a longer time period. A pressure of about 525 hPa was established in the pulpal space of 11 single specimens (n = 11) that were seated in a storage vessel filled with physiologic saline by increasing the pressure in the ADF reservoir to 500 hPa. When a continuous stream of ADF could be seen passing through the specimen window, the experiment was started (Figs. 2 and 3). Hitherto, the specimens were rinsed with physiologic saline from outside, gently dried and immediately put into a new centrifuge tube (tube 1) with 20 ml of fresh physiologic saline, and the time was measured beginning with this starting point (time = 0 min in Fig. 3). The specimens were kept in the tube for 30 min and then removed (at time point 30 min in Fig. 3) and stored transiently at reduced, environmental pressure conditions (=25 hPa of the apparatus) in a storage vessel also containing fresh physiologic saline. After 15 min of storage, the specimens were put into a new centrifuge tube (tube 2, at time point 45 min) with 20 ml fresh saline and the pressure was raised again to 525 hPa. After another 30 min (at time point 75 min), the specimens were removed from tube 2. These alternating pressure conditions were repeated five times (up to time point 255 min), and the absorption caused by the dye stained ADF in each centrifuge tube (tubes 1 to 6) was measured with the photometer at a wavelength of 664 nm. During perfusion time, the absence of ADF perfusion beyond the previously described window was determined visually. In case of any leakage beyond this window, e.g. through a gap between specimen holder and root, the respective specimen was discarded.Fig. 2

Bottom Line: In experiment 2, permeability was different within the specimens and decreased highly linearly with time, allowing the prediction of future values.In experiment 3, Seal&Protect® completely occluded dentinal tubules. elmex fluid® increased tubular permeability by about 30 % compared to control.The described in vitro model allows evaluation of potential agents for the treatment of DH at the clinically relevant cervical region of human teeth.

View Article: PubMed Central - PubMed

Affiliation: Department of Preventive Dentistry, Periodontology, and Cariology, University of Zürich Center for Dental Medicine, Plattenstrasse 11, 8032, Zürich, Switzerland, holger.jungbluth@zzm.uzh.ch.

ABSTRACT

Objectives: The aim of this series of studies was the development and validation of a new model for evaluation of dentinal hypersensitivity (DH) therapies.

Materials and methods: Roots from extracted human teeth were sealed with a flowable composite. In the cervical area, a 3-mm-wide circular window was ground through the seal 1 mm deep into dentine. The pulp lumen was connected to a reservoir of artificial dentinal fluid (ADF) containing protein, mineral salts and methylene blue. At increased pulpal pressure, the ADF released through the said window was collected in containers each with 20 ml of physiologic saline for a consecutive series of 30-min intervals and ADF concentration (absorption) was determined photometrically. The model was verified by three experiments. In experiment 1, the lower limit of quantification (LLoQ, coefficient of variation = 20 % and difference of 5 standard deviations (SD) from blank) of ADF in physiologic saline was determined by measuring the absorption of 15 dilutions of ADF in physiologic saline (containing 0.625 ng to 12.5 μg methylene blue/ml) photometrically for ten times. In experiment 2, long-term linearity of ADF perfusion/outflow was investigated using 11 specimens. The ADF released through the window was collected in the said containers separately for each consecutive interval of 30 min for up to 240 min. Absorption was determined and analysed by linear regression over time. In experiment 3, perfusion before (2×) and after single treatment according to the following three groups was measured: BisGMA-based sealant (Seal&Protect®), an acidic fluoride solution (elmex fluid®) and control (no treatment).

Results: In experiment 1, the LLoQ was 0.005 μg methylene blue/ml. In experiment 2, permeability was different within the specimens and decreased highly linearly with time, allowing the prediction of future values. In experiment 3, Seal&Protect® completely occluded dentinal tubules. elmex fluid® increased tubular permeability by about 30 % compared to control.

Conclusions: A model comprising the use of artificial dentinal fluid was developed and validated allowing screening of therapeutic agents for the treatment of DH through reliable measurement of permeability of cervical root dentine.

Clinical relevance: The described in vitro model allows evaluation of potential agents for the treatment of DH at the clinically relevant cervical region of human teeth.

No MeSH data available.


Related in: MedlinePlus