Objective

Improved methods are needed to remineralize dentin caries in order to promote conservation of dentin tissue and minimize the surgical interventions that are currently required for clinical treatment. Here, we test the hypothesis that bulk substrates can be effectively mineralized via a dual analog system proposed by others, using a tripolyphosphate (TPP) "templating analog" and a poly(acrylic acid) (PAA) or poly(aspartic acid) (pAsp) "sequestration analog," the latter of which generates the polymer-induced liquid-precursor (PILP) mineralization process studied in our laboratory.

Material & methods

Demineralized human dentin slices were remineralized with and without pre-treatment with TPP, using either PAA or pAsp as the PILP process-directing agent. A control experiment with no polymer present was used for comparison.

Results

No mineralization was observed in any of the PAA groups. In both the pAsp and no polymer groups, TPP inhibited mineralization on the surfaces of the specimens but promoted mineralization within the interiors. Pre-treatment with TPP enhanced overall mineralization of the pAsp group. However, when analysed via TEM, regions with little mineral were still present.

Conclusion

Poly(acrylic acid) was unable to remineralize demineralized dentin slices under the conditions employed, even when pre-treated with TPP. However, pre-treatment with TPP enhanced overall mineralization of specimens that were PILP-remineralized using pAsp.

Objective

Improved methods are needed to remineralize dentin caries in order to promote conservation of dentin tissue and minimize the surgical interventions that are currently required for clinical treatment. Here, we test the hypothesis that bulk substrates can be effectively mineralized via a dual analog system proposed by others, using a tripolyphosphate (TPP) "templating analog" and a poly(acrylic acid) (PAA) or poly(aspartic acid) (pAsp) "sequestration analog," the latter of which generates the polymer-induced liquid-precursor (PILP) mineralization process studied in our laboratory.

Material & methods

Demineralized human dentin slices were remineralized with and without pre-treatment with TPP, using either PAA or pAsp as the PILP process-directing agent. A control experiment with no polymer present was used for comparison.

Results

No mineralization was observed in any of the PAA groups. In both the pAsp and no polymer groups, TPP inhibited mineralization on the surfaces of the specimens but promoted mineralization within the interiors. Pre-treatment with TPP enhanced overall mineralization of the pAsp group. However, when analysed via TEM, regions with little mineral were still present.

Conclusion

Poly(acrylic acid) was unable to remineralize demineralized dentin slices under the conditions employed, even when pre-treated with TPP. However, pre-treatment with TPP enhanced overall mineralization of specimens that were PILP-remineralized using pAsp.

OBJECTIVE: The polymer-induced liquid-precursor (PILP) mineralization process has been shown to remineralize artificial dentin lesions to levels consistent with those of native dentin. However, nanoindentation revealed that the moduli of those remineralized lesions were only ∼50% that of native dentin. We hypothesize that this may be due to the PILP process having been previously optimized to obtain high amounts (∼70wt%) of intrafibrillar crystals, but without sufficient interfibrillar mineral, another significant component of dentin. METHODS: Fluoride was added to the PILP-mineralization of collagen from rat tail tendon at varying concentrations to determine if a better balance of intra- versus inter-fibrillar mineralization could be obtained, as determined by electron microscopy. Nanoindentation was used to determine if fluoridated apatite could improve the mechanical properties of the composites. RESULTS: Fluoride was successfully incorporated into the PILP-mineralization of rat tail tendon and resulted in collagen-mineral composite systems with the mineral phase of hydroxyapatite containing various levels of fluoridation. As the fluoride concentration increased, the crystals became larger and more rod-like, with an increasing tendency to form on the fibril surfaces rather than the interior. Nanomechanical testing of the mineralized tendons revealed that fluoride addition did not increase modulus over PILP mineralization alone. This likely resulted from the separated nature of collagen fibrils that comprise tendon, which does not provide lateral reinforcement and therefore may not be suited for the compressive loads of nanoindentation. SIGNIFICANCE: This work contributes to the development of minimally invasive approaches to caries treatment by determining if collagen can be functionally mineralized.

Objective

The polymer-induced liquid-precursor (PILP) mineralization process has been shown to remineralize artificial dentin lesions to levels consistent with those of native dentin. However, nanoindentation revealed that the moduli of those remineralized lesions were only ∼50% that of native dentin. We hypothesize that this may be due to the PILP process having been previously optimized to obtain high amounts (∼70wt%) of intrafibrillar crystals, but without sufficient interfibrillar mineral, another significant component of dentin.

Methods

Fluoride was added to the PILP-mineralization of collagen from rat tail tendon at varying concentrations to determine if a better balance of intra- versus inter-fibrillar mineralization could be obtained, as determined by electron microscopy. Nanoindentation was used to determine if fluoridated apatite could improve the mechanical properties of the composites.

Results

Fluoride was successfully incorporated into the PILP-mineralization of rat tail tendon and resulted in collagen-mineral composite systems with the mineral phase of hydroxyapatite containing various levels of fluoridation. As the fluoride concentration increased, the crystals became larger and more rod-like, with an increasing tendency to form on the fibril surfaces rather than the interior. Nanomechanical testing of the mineralized tendons revealed that fluoride addition did not increase modulus over PILP mineralization alone. This likely resulted from the separated nature of collagen fibrils that comprise tendon, which does not provide lateral reinforcement and therefore may not be suited for the compressive loads of nanoindentation.

Significance

This work contributes to the development of minimally invasive approaches to caries treatment by determining if collagen can be functionally mineralized.

The aim of this study was to evaluate the feasibility of applying the polymer-induced liquid-precursor (PILP) method to enhance silver diamine fluoride (SDF) therapy. One hundred forty micrometer deep artificial caries lesions were treated with (A) 38% SDF solution and (B) 38% SDF containing poly-L-aspartic acid (pASP). Changes in the nanomechanical profile across the lesion were evaluated. Hydrated artificial lesions had a low reduced elastic modulus (0.3 GPa) and nanohardness (0.02 GPa) region extending about 100 μm into the lesion, with a gradual linear increase to about 168 μm where the values plateaued to around 18 GPa/1.0 GPa. Topical application of SDF resulted in significantly recovered properties (p<0.001). SDF containing pASP resulted in greater nanomechanical properties compared to SDF alone, showing similar sloped regions up to 96 μm, then SDF alone dropped while SDF containing pASP continued at a modest slope until reaching normal at 144 μm. This nanoindentation study shows enhanced SDF therapy using the PILP method.

The addition of charged polymers, like poly-aspartic acid (pAsp), to mineralizing solutions allows for transport of calcium and phosphate ions into the lumen of collagen fibrils and subsequent crystallization of oriented apatite crystals by the so-called Polymer-Induced Liquid Precursor (PILP) mineralization process, leading to the functional recovery of artificial dentin lesions by intrafibrillar mineralization of collagen. OBJECTIVE:To evaluate the feasibility of applying the PILP method as part of a restorative treatment and test for effectiveness to functionally remineralize artificial lesions in dentin. MATERIALS AND METHODS:Two methods of providing pAsp to standardized artificial lesions during a restorative procedure were applied: (A) pAsp was mixed into commercial RMGI (resin modified glass ionomer) cement formulations and (B) pAsp was added at high concentration (25mg/ml) in solution to rehydrate lesions before restoring with a RMGI cement. All specimens were immersed in simulated body fluid for two weeks to allow for remineralization and then analyzed for dehydration shrinkage, integrity of cement-dentin interface, degree of mineralization, and changes in the nanomechanical profile (E-modulus) across the lesion. RESULTS:After the remineralization treatment, lesion shrinkage was significantly reduced for all treatment groups compared to demineralized samples. Pores developed in RMGI when pAsp was added. A thin layer at the dentin-cement interface, rich in polymer formed possibly from a reaction between pAsp and the RMGI. When analyzed by SEM under vacuum, most lesions delaminated from the cement interface. EDS-analysis showed some but not full recovery of calcium and phosphorous levels for treatment groups that involved pAsp. Nanoindentations placed across the interface indicated improvement for RMGI containing 40% pAsp, and were significantly elevated when lesions were rehydrated with pAsp before being restored with RMGI. In particular the most demineralized outer zone recovered substantially in the elastic modulus, suggesting that functional remineralization has been initiated by pAsp delivery upon rehydration of air-dried demineralized dentin. In contrast, the effectiveness of the RMGI on functional remineralization of dentin was minimal when pAsp was absent. SIGNIFICANCE:Incorporation of pAsp into restorative treatments using RMGIs promises to be a feasible way to induce the PILP-mineralization process in a clinical setting and to repair the structure and properties of dentin damaged by the caries process.

The addition of charged polymers, like poly-aspartic acid (pAsp), to mineralizing solutions allows for transport of calcium and phosphate ions into the lumen of collagen fibrils and subsequent crystallization of oriented apatite crystals by the so-called Polymer-Induced Liquid Precursor (PILP) mineralization process, leading to the functional recovery of artificial dentin lesions by intrafibrillar mineralization of collagen.

Objective

To evaluate the feasibility of applying the PILP method as part of a restorative treatment and test for effectiveness to functionally remineralize artificial lesions in dentin.

Materials and methods

Two methods of providing pAsp to standardized artificial lesions during a restorative procedure were applied: (A) pAsp was mixed into commercial RMGI (resin modified glass ionomer) cement formulations and (B) pAsp was added at high concentration (25mg/ml) in solution to rehydrate lesions before restoring with a RMGI cement. All specimens were immersed in simulated body fluid for two weeks to allow for remineralization and then analyzed for dehydration shrinkage, integrity of cement-dentin interface, degree of mineralization, and changes in the nanomechanical profile (E-modulus) across the lesion.

Results

After the remineralization treatment, lesion shrinkage was significantly reduced for all treatment groups compared to demineralized samples. Pores developed in RMGI when pAsp was added. A thin layer at the dentin-cement interface, rich in polymer formed possibly from a reaction between pAsp and the RMGI. When analyzed by SEM under vacuum, most lesions delaminated from the cement interface. EDS-analysis showed some but not full recovery of calcium and phosphorous levels for treatment groups that involved pAsp. Nanoindentations placed across the interface indicated improvement for RMGI containing 40% pAsp, and were significantly elevated when lesions were rehydrated with pAsp before being restored with RMGI. In particular the most demineralized outer zone recovered substantially in the elastic modulus, suggesting that functional remineralization has been initiated by pAsp delivery upon rehydration of air-dried demineralized dentin. In contrast, the effectiveness of the RMGI on functional remineralization of dentin was minimal when pAsp was absent.

Significance

Incorporation of pAsp into restorative treatments using RMGIs promises to be a feasible way to induce the PILP-mineralization process in a clinical setting and to repair the structure and properties of dentin damaged by the caries process.

It was hypothesized that applying the polymer-induced liquid-precursor (PILP) system to artificial lesions would result in time-dependent functional remineralization of carious dentin lesions that restores the mechanical properties of demineralized dentin matrix. 140 µm deep artificial caries lesions were remineralized via the PILP process for 7-28 days at 37°C to determine temporal remineralization characteristics. Poly-L-aspartic acid (27 KDa) was used as the polymeric process-directing agent and was added to the remineralization solution at a calcium-to-phosphate ratio of 2.14 (mol/mol). Nanomechanical properties of hydrated artificial lesions had a low reduced elastic modulus (E(R) = 0.2 GPa) region extending about 70 μm into the lesion, with a sloped region to about 140 μm where values reached normal dentin (18-20 GPa). After 7 days specimens recovered mechanical properties in the sloped region by 51% compared to the artificial lesion. Between 7-14 days, recovery of the outer portion of the lesion continued to a level of about 10 GPa with 74% improvement. 28 days of PILP mineralization resulted in 91% improvement of E(R) compared to the artificial lesion. These differences were statistically significant as determined from change-point diagrams. Mineral profiles determined by micro x-ray computed tomography were shallower than those determined by nanoindentation, and showed similar changes over time, but full mineral recovery occurred after 14 days in both the outer and sloped portions of the lesion. Scanning electron microscopy and energy dispersive x-ray analysis showed similar morphologies that were distinct from normal dentin with a clear line of demarcation between the outer and sloped portions of the lesion. Transmission electron microscopy and selected area electron diffraction showed that the starting lesions contained some residual mineral in the outer portions, which exhibited poor crystallinity. During remineralization, intrafibrillar mineral increased and crystallinity improved with intrafibrillar mineral exhibiting the orientation found in normal dentin or bone.