Heinz Duschner

Influence of bisphosphonates on endothelial cells, fibroblasts, and osteogenic cells

Bisphosphonate-associated osteonecrosis of the jaws (BP-ONJ) is a side effect primarily in patients receiving highly potent nitrogen-containing bisphosphonates. The exact etiopathology is unknown. In addition to reduced bone remodeling, there may also be an impact on soft tissues. The impact of nitrogen- (ibandronate, pamidronate, zoledronate) and non-nitrogen-containing bisphosphonates (clodronate) on human umbilicord vein endothelial cells (HUVEC), fibroblasts and osteogenic cells was analyzed employing cell viability testing and a scratch wound assay. The impact on the cell morphology of vital-stained osteogenic cells was investigated by cell visualization (confocal laser scanning microscopy). Pamidronate and zoledronate had the greatest negative impact on all cell lines, whereas the impact of ibandronate and clodronate was less distinct. The effect of clodronate on HUVEC and fibroblasts was particularly marginal. BP-ONJ could be a multifactorial event with multicellular impairments. This might result in altered wound healing. The increased impact of the highly potent bisphosphonates, particularly on non-bone cells, may explain the higher occurrence of BP-ONJ.

Interactions between cells and titanium surfaces

The interaction between cells and implant materials is determined by the surface structure and/or surface composition of the material. In the past years, titanium and titanium alloys have proved their superiority over other implant materials in many clinical applications. This predominant behaviour is caused by a dense passive oxide layer which forms within milliseconds in oxidizing media. Titanium dioxide layers of 100 nm thickness were produced on the surface of cp-titanium grade 2, and on an experimental alloy of high vanadium content (Ti1.5Al25V) as a harmful control. The layers were produced by thermal and anodic oxidation and by coating by means of the sol-gel process. The resulting oxide layers were characterized with respect of their structure and chemical composition. In cell tests (proliferation, MTT, morphology, actin staining), the reaction of the cells was examined. It was shown that the sol-gel-produced titanium oxide layer is able to shield the cells from toxic alloying elements, with the result that the cell reaction is influenced only by the thin titanium oxide surface layer and not by the composition of the bulk material.

Structural Changes in Human Dental Enamel after Subablative Erbium Laser Irradiation and Its Potential Use for Caries Prevention

Numerous studies have confirmed the potential of erbium laser irradiation for increasing the acid resistance of dental enamel. The objective of the present paper was to investigate the effect of subablative erbium laser irradiation on the structure and acid resistance of dental enamel by means of confocal laser scanning microscopy (CLSM). To this end, 12 samples of human dental enamel were irradiated with subablative energy densities (Φ) of an Er:YAG (λ = 2.94 µm, Φ = 6 J/cm2) and an Er:YSGG laser (λ = 2.79 µm, Φ = 8 J/cm2). The enamel surfaces of 6 samples were polished prior to irradiation. The remaining 6 samples were left intact (without polishing procedures) and, in the further course of the study, they were subjected to 1-week in situ demineralisation. All irradiated test surfaces were assigned a control surface on the same sample. The changes following laser irradiation and the in situ wearing time were assessed qualitatively using a confocal laser scanning microscope. The irradiation of dental enamel with subablative erbium laser irradiation produces fine cracks in the enamel surface. These cracks act as starting points for acid attack and favour deep demineralisation. These changes reduce or eliminate the positive effect of subablative erbium laser irradiation observed in connection with caries-preventive use. The clinical use of subablative erbium laser irradiation to prevent caries would appear not to make sense under the conditions studied.

Effect of diode laser irradiation on root surfaces in vitro.

OBJECTIVE The objective of this study was to evaluate possible morphological alterations of root surfaces after GaAlAs-diode laser (809 nm) irradiation under standardized in vitro conditions. MATERIALS AND METHODS Root specimens obtained from extracted periodontally diseased teeth were scaled and root planed with curettes followed by air-powder abrasive treatment prior to lasing. The variable parameters were power output (0.5-2.5 W) and exposure time (10-30 sec per specimen). Additionally, the effect of a saline solution and a human blood film on the root surface was investigated. The root segments were analyzed by means of a reflected light microscope. Photographs before and after irradiation were taken and evaluated. The scale of carbonization was quantified using a grid laid over the photographs. Specimens with distinct morphological changes were analyzed with a scanning electron microscope. RESULTS Lasing dry specimens and specimens moistened with saline resulted in no detectable alterations, irrespective of irradiation time and power output applied. Depending on different settings, irradiation caused severe damages to the root surface when segments were covered by a thin blood film. Irradiation at a power output of 1 Watt and below had barely any negative effect on the root surface, whereas lasing at 1.5, 2.0, and 2.5 Watt resulted in partial or total carbonizations of the root samples. The angle of irradiation had a significant effect on the scale of the root surface damage (Mann-Whitney U test,p < 0.05). CONCLUSION The diode laser may cause damage to periodontal hard tissues if irradiation parameters are not adequate.

Long‐term response of osteogenic cells on micron and submicron‐scale‐structured hydrophilic titanium surfaces: sequence of cell proliferation and cell differentiation

OBJECTIVE Modifications of surface topography and surface chemistry are key factors for guiding target cells during dental implant healing. Recent in vitro studies confirmed promotion of early osteogenic cell differentiation on submicron scaled surfaces in particular when hydrophilized. However, no long-term observations on both osteogenic cell proliferation as well as on cell maturation have been reported for respectively modified surfaces. Aim of this study was to monitor osteogenic cell proliferation and expression of specific osteogenic cell differentiation markers on a protein level over an extended period of 3 weeks with respect to surface modifications. MATERIAL AND METHODS Modified titanium (Ti) disks were obtained from Institute Straumann, representing the following surfaces: smooth pretreatment (PT), sandblasted/acid etched (SLA), and hydrophilized (modSLA). Surface topography was analyzed by scanning electron microscopy, surface elemental composition was assessed by X-Ray Photoelectronic Spectroscopy (XPS). Tissue culture polystyrene (TCPS) served as a control substrate. Human osteogenic cells (HOB-c) were cultivated on the respective substrates. After 24 hrs, 48 hrs, 72 hrs, 7 d, 14 d and 21 d, cell count was assessed as well as osteogenic cell differentiation utilizing cellular Quantitative Immuno-Cytochemistry (QIC) assay for collagen type I (COL), alkaline phosphatase (AP), osteopontin (OPN) and osteocalcin (OC). Data were normalized with respect to internal controls. RESULTS In contrast to the other modified Ti disks, modSLA stands out due to low surface carbon contamination. TCPS and PT surfaces preserved a rather immature, mitotic active osteogenic phenotype (high proliferation rates, no increase of OC production), SLA and especially modSLA surfaces promoted the maturation of osteogenic precursors into post-mitotic osteoblasts. In detail, modSLA resulted in lowest cell proliferation rates, but exhibited highest expression rates of OC. DISCUSSION Our results, which confirm previous studies, reveal long-term promotion of osteogenic cell maturation by topography (micron and submicron scale roughness) and surface hydrophilicity.

Mixed silane self assembled monolayers and their in situ modification

Mixed self assembled silane monolayers were prepared by coadsorption of Br and methyl terminated trichlorosilanes. By subsequent in situ modification the Br groups were converted into amino groups which can be used to tether polyglutamates to the surface. Here we describe the characterization of the monolayers by means of contact angle measurements and X-ray photoelectron spectroscopy (XPS). Both methods clearly indicate that the Br terminated silane is preferentially adsorbed from the solution. Following the in situ modification by XPS a yield of approximately 80% was determined for the substitution of the bromine by azide, whereas the yield for subsequent reduction to the amine is close to 100%.

Using ultrasound transmission velocity to analyse the mechanical properties of teeth after in vitro, in situ, and in vivo irradiation.

Abstract Ultrasound transmission velocity (UTV) in isotropic material as a measure for the modulus of elasticity was correlated to mechanical properties. Changes in micromechanical properties of radiated teeth and influence of the oral cavity were to be evaluated nondestructively. UTV was measured in extracted teeth after 36 Gy and 62 Gy of in situ (enorally, with no contact to the oral cavity) and in vitro irradiation. Relative to controls, teeth subjected to 62 Gy in vivo showed higher UTV values for dentine and enamel. Sound teeth irradiated with 60 Gy in situ also showed higher UTV values for enamel, whereas dentine values were not significantly different from those of control. The mechanical properties of teeth irradiated in vitro were affected only after high experimental doses of up to 500 Gy. The difference between in vivo and in vitro mechanical properties may be due to radioxerostomia-induced damages as well as the status of dentine vitality. This supports the concept of direct radiation-induced damage in synergy with radioxerostomia-induced caries.

Caries Development in Fluoridated and Non-Fluoridated Deciduous and Permanent Enamel in Situ Examined by Microradiography and Confocal Laser Scanning Microscopy

The aim of the study was to compare initial caries development in fluoridated and non-fluoridated deciduous and permanent enamel in situ. Enamel slabs were mounted in removable appliances and worn for 4 wks. Significantly larger lesions developed in deciduous than in permanent enamel when no topical fluorides were used. Fluoride mouthrinsing partly prevented lesion development in deciduous and completely in permanent enamel. Initial enamel caries not detected by microradiography can be visualized by CLSM (confocal laser scanning microscopy).

Bestimmung der Größe der spezifischen Oberfläche von Knochenersatzmaterialien mittels Gasadsorption

Fragestellung: Die Oberfläche sowie die Mikro- und Mesoporosität von Knochenersatzmaterialien beeinflussen deren chemische und biologische Eigenschaften. Daher wurden in dieser Untersuchung die Größe der spezifischen Oberfläche sowie die Verteilung der Porendurchmesser (Poren < 1 μm) in Knochenersatzmaterialien bestimmt. Material und Methode: Die untersuchten Hydroxylapatite waren synthetischen, bovinen und phytotrophen Ursprungs. Die Trikalziumphosphate und die Biogläser umfassten nur rein synthetische Materialien. ¶Die Gasadsorptionsuntersuchung je einer Probe erfolgte mit Hilfe eines volumetrischen N 2 -Kr-Systems (ASAP 2010, Micromeritics). Zusätzlich wurde für Materialien mit spezifischen Oberflächen ¶> 2 m 2 /g die Porengrößenverteilung nach der BJH-Methode ermittelt. Ergebnisse: 1. Spezifische Oberfläche: 2 der Materialien zeigten eine auffallend große spezifische Oberfläche (BioOss 79,7 m 2 /g, Algipore neu 14,6 m 2 /g). Eine mittlere Oberfläche zeigten Algipore alt (4,9 m 2 /g) und Interpore 200 (2,64 m 2 /g). Die übrigen Materialien zeigten nur kleine Oberflächen (Ceros 80 1,8 m 2 /g, Ceros 82 ¶1,31 m 2 /g, Cerasorb 1,2 m 2 /g, Biobase 0,7 m 2 /g, Endobone 0,7 m 2 /g, Perioglas 0,6 m 2 /g, Allotropat 50 0,2 m 2 /g, Biogran 0,2 m 2 /g). 2. Häufigkeitsverteilung des Porendurchmessers: Die Materialien mit großen und mittleren spezifischen Oberflächen zeigten folgende Porendurchmesser: BioOss 2–50 nm, Algipore neu 2–¶100 nm, Algipore alt 5–50 nm, Interpore 200 2–100 nm. Porengrößen < 2 nm fanden sich kaum. Schlussfolgerungen: Die Materialien BioOss, Algipore alt und neu und Interpore 200 haben ein großes interkonnektierendes Mesoporensystem (Durchmesser < 1 μm). Für die Materialien Biobase, Endobone, Perioglas, Allotropat 50 und Biogran ist dies nicht anzunehmen. Die Materialien Ceros 80, Ceros 82 und Cerasorb zeigen eine dazwischen liegende spezifische Oberflächengröße und weisen einen mäßigen Anteil von solchen interkonnektierenden Poren auf. Ein Einfluss der interkonnektierenden Porosität und der deutlich unterschiedlichen spezifischen Oberflächen auf das Verhalten der Knochenersatzmaterialien in vivo ist nahe liegend. The surface area and the microporosity of bone regeneration materials influence their chemical and ¶biological properties. Therefore, the size of the specific surface area and the distribution of the pore diameters (pores < 1 μm) of bone regeneration materials were analyzed within this study. The analyzed hydroxyapatites were of synthetic, bovine, and phytotroph origin. The tricalcium phosphates and the bioglasses included only synthetic materials. The gas adsorption of each specimen was analyzed using a volumetric N2/Kr system (ASAP 2010, Micromeritics). Additionally, for materials with a specific surface area (> 2 m2/g) the pore size distribution was evaluated by the BJH-method. Two of the ¶materials evaluated astonishingly large dimensions of the specific surface area (BioOss 79.7 m2/g, Algipore new 14.6 m2/g). A medium ¶surface area was found for Algipore old (4.9 m2/g) and Interpore200 ¶(2.64 m2/g). All other included materials showed only small sizes of the specific surface area (Ceros80 ¶1.8 m2/g, Ceros82 1.31 m2/g, Cerasorb 1.2 m2/g, Biobase 0.7 m2/g, Endobone 0.7 m2/g, Perioglas 0.6 m2/g, Allotropat50 0.23 m2/g, Biogran ¶0.2 m2/g). The materials with large and medium sizes of the specific surface area evaluated the following pore diameters: BioOss 2–50 nm, Algipore new 2–100 nm, Algipore old 5–50 nm, Interpore200 2–100 nm. Pore sizes less than 2 nm were not found in relevant numbers. The materials BioOss, old and new Algipore, and Interpore200 contain a large interconnecting mesopore system (diameter < 1 μm). For the materials Biobase, Endobone, Perioglas, Allotropat 50, and Biogran this cannot be assumed. The materials Ceros80, Ceros82, and Cerasorb evaluated a specific surface area between those and might include only a small part of these interconnecting pores. An influence of the interconnecting porosity and the different sizes of the specific surface areas on the biological behavior of the bone regeneration materials can be suggested.

Neue Erkenntnisse zur Ätiologie der sogenannten Strahlenkaries

ZusammenfassungFragestellungDie Ätiologie der Strahlenkaries wird trotz vieler Untersuchungen kontrovers diskutiert: Die These einer direkten radiogenen Läsion des Zahnhartgewebes steht der einer indirekten Pathogenese über die Radioxerostomie gegenüber.MethodeIn einer systematischen Studie wurden Zähne mit manifester Strahlenkaries (Gruppe 1, zirka 60 Gy, langes Intervall bis zur Extraktion) und klinisch kariesfreie Zähne (Gruppe 2, zirka 30 Gy, kurzes Intervall) Zahnproben nach experimenteller enoraler In-situ- (60 Gy; Gruppe 3) und In-vitro-Bestrahlung (500 bis 2500 Gy; Gruppe 4) gegenübergestellt (jeweils60Co-Bestrahlung). Diese Proben wurden mit gesunden Zähnen als Kontrollen (Gruppe 5) verglichen. Durch konfokale Laser-Scanning-Mikroskopie (CLSM) erfolgten zerstörungsfreie Histotomogramme an frischen Längsschliffen und an Technovit-eingebetteten Schliffen (Trenn-Dünnschliff-Methode) mit Darstellung der Schmelz-Dentin-Grenze.ErgebnisseDrei charakteristische Veränderungen traten nur an Zahnproben von Radiotherapiepatienten auf: 1. Die grenznahen Aufzweigungen (Ramifikationen) der Odontoblastenfortsätze waren rarefiziert, 2. die Dentinkanälchen erreichten die Hartgewebsgrenze nicht mehr und 3. zeigte sich grenznah eine zirka 10 μm breite Zone geringerer Intensität remittierten Lichtes im Dentin.SchlußfolgerungenDie Obliteration der Dentinkanälchen, der eine Degeneration der Odontoblastenfortsätze vorausgeht, ist offensichtlich das Ergebnis eines direkten radiogenen Zellschadens mit Vaskularisations- und Stoffwechseleinschränkung im Bereich der Endigungen der Odontoblasten („letzte Wiese”). Das Stoffwechseldefizit vermittelt mit einer Latenz einen Parenchymschaden (hyporemittierende Zone), der die funktionellen Symptome (unterminierende Karies) erklärt. Die mikromorphologische Manifestation dieses direkten Strahlenschadens setzt den vitalen Zahn voraus und kann deshalb in situ und in vitro nicht beobachtet werden.AbstractAimIn spite of a great number of relevant studies the etiology of radiation induced caries still is discussed in controversy: The assumption of direct radiation induced lesions in the hard tissue is in contrast to an indirect pathogenesis mediated via radio-xerostomia.MethodsA systematic study is presented, comparing teeth with a manifest radiation caries (group 1, about 60 Gy, long interval to the extraction) and clinically caries free teeth (group 2, about 30 Gy, short interval) with tooth specimens after an experimental enoral (in situ) irradiation (60 Gy, group 3) and after in vitro irradiation (500 to 2.500 Gy, group 4).60Co was the irradiation source. Sound teeth were used as a standard (group 5). For non destructive visualisation of subsurface histotomograms by confocal laser scanning microscopy (CLSM) teeth were either used as fresh sections or as Technovit embedded thin slices (sawing grinding technique).ResultsTooth samples from radiotherapy patients (cancer therapeutic doses, long interval before extraction; group 1) showed three characteristic changes: 1. rarefication of the branching (ramification) of odontoblast processes near the junction, 2. dentine tubules end infront of the interface to the hard tissue and 3. in dentine the interface is characterised by an zone (about 10 μm wide) of low intensity of the remitted light.ConclusionsThe obliteration of the dentine tubules, preceded by a degeneration of the odontoblast processes, is obviously the result of a direct radiogenic cell damage with hampered vascularisation and metabolism particularly in the area of the terminations of the odontoblast processes. The deficit in metabolism combined with a latent damage of the parenchyma (hypo-remitting zone) is evidence for the functional symptoms (subsurface caries). The prerequisite for the micromorphological manifestation of this direct irradiation damage is a vital tooth and in consequence cannot be simulated in situ or in vivo.AIM In spite of a great number of relevant studies the etiology of radiation induced caries still is discussed in controversy: The assumption of direct radiation induced lesions in the hard tissue is in contrast to an indirect pathogenesis mediated via radio-xerostomia. METHODS A systematic study is presented, comparing teeth with a manifest radiation caries (group 1, about 60 Gy, long interval to the extraction) and clinically caries free teeth (group 2, about 30 Gy, short interval) with tooth specimens after an experimental enoral (in situ) irradiation (60 Gy, group 3) and after in vitro irradiation (500 to 2,500 Gy, group 4). 60Co was the irradiation source. Sound teeth were used as a standard (group 5). For non destructive visualisation of subsurface histotomograms by confocal laser scanning microscopy (CLSM) teeth were either used as fresh sections or as Technovit embedded thin slices (sawing grinding technique). RESULTS Tooth samples from radiotherapy patients (cancer therapeutic doses, long interval before extraction; group 1) showed three characteristic changes: 1, rarefcation of the branching (ramification) of odontoblast processes near the junction, 2. dentine tubules end in front of the interface to the hard tissue and 3, in dentine the interface is characterised by an zone (about 10 microns wide) of low intensity of the remitted light. CONCLUSIONS The obliteration of the dentine tubules, preceded by a degeneration of the odontoblast processes, is obviously the result of a direct radiogenic cell damage with hampered vascularisation and metabolism particularly in the area of the terminations of the odontoblast processes. The deficit in metabolism combined with a latent damage of the parenchyma (hypo-remitting zone) is evidence for the functional symptoms (subsurface caries). The prerequisite for the micromorphological manifestation of this direct irradiation damage is a vital tooth and in consequence cannot be simulated in situ or in vivo.