Morphological characterization using scanning electron microscopy of fly artifacts deposited by Calliphora vomitoria (Diptera: Calliphoridae) on household materials.

Insects found at a crime scene can produce traces referred to as fly artifacts (FA) due to their movement over the corpse and the manner in which they feed upon it. These can be detrimental for carrying out criminal investigations. Confusing a FA with a genuine bloodspot can lead to misinterpretations, also taking into consideration that FA may contain a human DNA profile. The aim of the present study was to employ scanning electron microscopy (SEM) for the analysis of FA produced by Calliphora vomitoria on hard surfaces and fabrics that are commonly present at crime scenes. FA and control bloodstains were produced under experimental conditions on metal, glass, plaster, cotton, and polyester. After macroscopic analysis, FA were examined at standard low (20-40 ×), medium low (300-600 ×), and high ultrastructural (1200 ×) magnification through a SEM Stereoscan 360, Leica, Cambridge. SEM analysis enabled the identification of distinctive features of FA on hard surfaces, namely, amorphous crystals, micro-crystals with a morphology similar to those of uric or micro-crystals with a comparable morphology to cholesterol, absent in controls. Moreover, red blood cells (RBC) were absent in FA but were always present in controls. On cotton, for both FA and controls, the drop was almost completely absorbed and thus indistinguishable from the underlying fabric texture. On polyester, FA showed amorphous/crystal-like deposits and no RBC, as observed on hard surfaces, except for those showing a completely flat surface. SEM analysis appeared to be suitable for differential diagnosis between FA and genuine bloodstains on hard surfaces, although the results may be inconclusive on tested fabrics.

Scanning electron microscopy in the identification of fly artifacts.

Bloodstain pattern analysis has a key role in crime scene reconstruction; however, it can be hampered by diverse confounding factors, such as insect activity which may lead to the production of small artifactual bloodstains, commonly referred to as fly artifacts (FA). Although several techniques aimed at distinguishing human bloodstains and FA have been developed, actually, no standardized and reproducible methodology is available. The aim of our study was to test the use of scanning electron microscopy (SEM) to distinguish human bloodstains from FA produced by Sarcophaga carnaria. FA and bloodstains have been produced on five different deposition surfaces under experimental conditions. After visual analysis, bloodstains and FA were analyzed at standard low (× 40-× 300) and high (× 600-× 1200) magnification through a Philips SEM 515. Although differential diagnosis between bloodstains and FA resulted often inconclusive at visual analysis, SEM analysis allowed the identification of additional key distinctive morphological features. In particular, on the surface of FA, small crystal-like and/or amorphous material deposits were observed. Such deposits were absent on bloodstains which, on the other hand, displayed red blood cells stacked in "rouleaux." Basing on these results and under our experimental conditions, SEM analysis resulted suitable to perform a differential diagnosis between bloodstains and FA produced from the insect activity of Sarcophaga carnaria.

Visualizing the Supramolecular Assembly of Collagen.

Among biological macromolecules, collagen enjoys quite a peculiar status. Making up as much as a third of the protein fraction of the body it is the main responsible for the functional properties of the extracellular matrix, which can be efficiently tuned and tailored by modifying the length, volume fraction, and spatial layout of its collagen content. The supramolecular aggregates of collagen are therefore subject to be investigation by several viewpoints and at different scales, from the finest interactions of individual collagen molecules to the spatial layout of fibril bundles. As a consequence, no treatise can pretend to be exhaustive about the several techniques that can be useful in different moments and/or for different purposes. So, in this chapter, we focus only on some applications of the transmission electron microscope (TEM), of the scanning electron microscope (SEM), and of the atomic force microscope (AFM).

Osteoclast differentiation from human blood precursors on biomimetic calcium-phosphate substrates.

The design of synthetic bone grafts to foster bone formation is a challenge in regenerative medicine. Understanding the interaction of bone substitutes with osteoclasts is essential, since osteoclasts not only drive a timely resorption of the biomaterial, but also trigger osteoblast activity. In this study, the adhesion and differentiation of human blood-derived osteoclast precursors (OCP) on two different micro-nanostructured biomimetic hydroxyapatite materials consisting in coarse (HA-C) and fine HA (HA-F) crystals, in comparison with sintered stoichiometric HA (sin-HA, reference material), were investigated. Osteoclasts were induced to differentiate by RANKL-containing supernatant using cell/substrate direct and indirect contact systems, and calcium (Ca++) and phosphorus (P5+) in culture medium were measured. We observed that OCP adhered to the experimental surfaces, and that osteoclast-like cells formed at a rate influenced by the micro- and nano-structure of HA, which also modulate extracellular Ca++. Qualitative differences were found between OCP on biomimetic HA-C and HA-F and their counterparts on plastic and sin-HA. On HA-C and HA-F cells shared typical features of mature osteoclasts, i.e. podosomes, multinuclearity, tartrate acid phosphatase (TRAP)-positive staining, and TRAP5b-enzyme release. However, cells were less in number compared to those on plastic or on sin-HA, and they did not express some specific osteoclast markers. In conclusion, blood-derived OCP are able to attach to biomimetic and sintered HA substrates, but their subsequent fusion and resorptive activity are hampered by surface micro-nano-structure. Indirect cultures suggest that fusion of OCP is sensitive to topography and to extracellular calcium. STATEMENT OF SIGNIFICANCE: The novelty of the paper is the differentiation of human blood-derived osteoclast precursors, instead of mouse-derived macrophages as used in most studies, directly on biomimetic micro-nano structured HA-based surfaces, as triggered by osteoblast-produced factors (RANKL/OPG), and influenced by chemistry and topography of the substrate(s). Biomimetic HA-surfaces, like those obtained in calcium phosphate cements, are very different from the conventional calcium phosphate ceramics, both in terms of topography and ion exchange. The role of these factors in modulating precursors' differentiation and activity is analysed. The system is closely reproducing the physiological process of attachment of host cells and further maturation to osteoclasts toward resorption of the substrate, which occurs in vivo after filling bone defects with the calcium phosphate grafts.

Differential proteomic and phenotypic behaviour of papillary and anaplastic thyroid cell lines.

Thyroid carcinomas account for a minority of all malignant tumours but, after those of the gonads, they represent the most common forms of endocrine cancers. They include several types, among which the papillary thyroid cancer (PTC) and the anaplastic thyroid cancer (ATC) are the best known. The two hystotypes display significant biological and clinical differences: PTC is a well differentiated form of tumour with a high incidence and a good prognosis, while the ATC is less frequent but represents one of the most aggressive endocrine tumours with morphological features of an undifferentiated type. To date, as far as we know, no conclusive studies, useful to design arrays of molecular markers, have been published illustrating the phenotypic and proteomic differences between these two tumours. The aim of this work was to perform a comparative analysis of two thyroid cancer cell lines, derived respectively from papillary (BCPAP) and anaplastic (8505C) thyroid carcinomas. The comparative analysis included cell behaviour assays and proteomic analysis by 2D-PAGE and mass spectrometry. The results have highlighted a new proteomic signature for the anaplastic carcinoma-derived cells, consistent with their high proliferation rate, motility propensity and metabolic shift, in relation to the well-differentiated PTC cells.

Early healing events around titanium implant devices with different surface microtopography: a pilot study in an in vivo rabbit model.

In the present pilot study, the authors morphologically investigated sandblasted, acid-etched surfaces (SLA) at very early experimental times. The tested devices were titanium plate-like implants with flattened wide lateral sides and jagged narrow sides. Because of these implant shape and placement site, the device gained a firm mechanical stability but the largest portion of the implant surface lacked direct contact with host bone and faced a wide peri-implant space rich in marrow tissue, intentionally created in order to study the interfacial interaction between metal surface and biological microenvironment. The insertion of titanium devices into the proximal tibia elicited a sequence of healing events. Newly formed bone proceeded through an early distance osteogenesis, common to both surfaces, and a delayed contact osteogenesis which seemed to follow different patterns at the two surfaces. In fact, SLA devices showed a more osteoconductive behavior retaining a less dense blood clot, which might be earlier and more easily replaced, and leading to a surface-conditioning layer which promotes osteogenic cell differentiation and appositional new bone deposition at the titanium surface. This model system is expected to provide a starting point for further investigations which clarify the early cellular and biomolecular events occurring at the metal surface.

Enhancing osteoconduction of PLLA-based nanocomposite scaffolds for bone regeneration using different biomimetic signals to MSCs.

In bone engineering, the adhesion, proliferation and differentiation of mesenchymal stromal cells rely on signaling from chemico-physical structure of the substrate, therefore prompting the design of mimetic "extracellular matrix"-like scaffolds. In this study, three-dimensional porous poly-L-lactic acid (PLLA)-based scaffolds have been mixed with different components, including single walled carbon nanotubes (CNT), micro-hydroxyapatite particles (HA), and BMP2, and treated with plasma (PT), to obtain four different nanocomposites: PLLA + CNT, PLLA + CNTHA, PLLA + CNT + HA + BMP2 and PLLA + CNT + HA + PT. Adult bone marrow mesenchymal stromal cells (MSCs) were derived from the femur of orthopaedic patients, seeded on the scaffolds and cultured under osteogenic induction up to differentiation and mineralization. The release of specific metabolites and temporal gene expression profiles of marrow-derived osteoprogenitors were analyzed at definite time points, relevant to in vitro culture as well as in vivo differentiation. As a result, the role of the different biomimetic components added to the PLLA matrix was deciphered, with BMP2-added scaffolds showing the highest biomimetic activity on cells differentiating to mature osteoblasts. The modification of a polymeric scaffold with reinforcing components which also work as biomimetic cues for cells can effectively direct osteoprogenitor cells differentiation, so as to shorten the time required for mineralization.

Evidence of a discrete axial structure in unimodal collagen fibrils.

The collagen fibrils of cornea, blood vessel walls, skin, gut, interstitial tissues, the sheath of tendons and nerves, and other connective tissues are known to be made of helically wound subfibrils winding at a constant angle to the fibril axis. A critical aspect of this model is that it requires the axial microfibrils to warp in an implausible way. This architecture lends itself quite naturally to an epitaxial layout where collagen microfibrils envelop a central core of a different nature. Here we demonstrate an axial domain in collagen fibrils from rabbit nerve sheath and tendon sheath by means of transmission electron microscopy after a histochemical reaction designed to evidence all polysaccharides and by tapping-mode atomic force microscopy. This axial domain was consistently found in fibrils with helical microfibrils but was not observed in tendon, whose microfibrils run longitudinal and parallel.

Correlative microscopy of bone in implant osteointegration studies.

Routine morphological analyses usually include investigations by light microscopy (LM), scanning electron microscopy (SEM), and transmission electron microscopy (TEM). Each of these techniques provides specific information on tissue morphology and all the obtained results are then combined to give an in-depth morphological overview of the examined sample. The limitations of this traditional comparative microscopy lie in the fact that each technique requires a different experimental sample, so that many specimens are necessary and the combined results come from different samples. The present study describes a technical procedure of correlative microscopy, which allows us to examine the same bone section first by LM and then, after appropriate processing, by SEM or TEM. Thanks to the possibility of analyzing the same undecalcified bone sections both by LM and SEM, the approach described in the present study allows us to make very accurate evaluations of old/new bone morphology at the bone-implant interface.

Contribution of glycosaminoglycans to the microstructural integrity of fibrillar and fiber crimps in tendons and ligaments.

The biomechanical roles of both tendons and ligaments are fulfilled by the extracellular matrix of these tissues. In particular, tension is mainly transmitted and resisted by protein (collagen, elastin) fibers, whereas compression is opposed by water-soluble glycosaminoglycans (GAGs). GAGs spanning the interfibrillar spaces and interacting with fibrils through the interfibrillar proteoglycans also seem to play a part in transmitting and resisting tensile stresses. Both tendons and ligaments showing similar composition, but different functional roles and collagen array, exhibit periodic undulations of collagen fibers or crimps. Each crimp is composed of many knots of each single fibril or fibrillar crimps. Fibrillar and fiber crimps play a mechanical role in absorbing the initial loading during elongation of both tendons and ligaments, and in recoiling fibrils and fibers when tissues have to return to their original length. This study investigated whether GAGs covalently attached to proteoglycan core proteins directly affect the 3D microstructural integrity of fibrillar crimp regions and fiber crimps in both tendons and ligaments. Achilles tendons and medial collateral ligaments of the knee from eight female Sprague-Dawley rats (90 days old) incubated in a chondroitinase ABC solution to remove GAGs were observed under a scanning electron microscope (SEM). In addition, isolated fibrils of these tissues obtained by mechanical disruption were analyzed by a transmission electron microscope (TEM). Both Achilles tendons and medial collateral ligaments of the rats after chemical or mechanical removal of GAGs still showed crimps and fibrillar crimps comparable to tissues with a normal GAG content. All fibrils in the fibrillar crimp region always twisted leftwards, thus changing their running plane, and then sharply bent, changing their course on a new plane. These data suggest that GAGs do not affect structural integrity or fibrillar crimp functions that seem mainly related to the local fibril leftward twisting and the alternating handedness of collagen from a molecular to a supramolecular level.

Influence of a zirconia sandblasting treated surface on peri-implant bone healing: An experimental study in sheep.

A sandblasting process with round zirconia (ZrO(2)) particles might be an alternative surface treatment to enhance the osseointegration of titanium dental implants. Our previous study on sheep compared smooth surface titanium implants (control) with implant surfaces sandblasted with two different granulations of ZrO(2). As the sandblasted surfaces proved superior, the present study further compared the ZrO(2) surface implant with other surface treatments currently employed: machined titanium (control), titanium oxide plasma sprayed (TPS) and alumina sandblasted (Al-SL) at different times after insertion (2, 4 and 12weeks). Twelve sheep were divided into three groups of four animals each and underwent implant insertion in tibia cortical bone under general anaesthesia. The implants with surrounding tissues were subjected to histology, histomorphometry, scanning electron microscopy and microhardness tests. The experimentation indicated that at 2weeks Zr-SL implants had the highest significant bone ingrowth (p<0.05) compared to the other implant surfaces, and a microhardness of newly formed bone inside the threads significantly higher than that of Ti. The present work shows that the ZrO(2) treatment produces better results in peri-implant newly formed bone than Ti and TPS processing, whereas its performance is similar to the Al-SL surface treatment.

Histological and ultrastructural evaluation of Leeds-Keio ligament 20 years after implant: a case report.

We were capable of undertaking a histological and ultrastructural evaluation of an intact Leeds-Keio ligament implanted 20 years ago to assess the neoligamentization process inside this artificial ligament. The histological evaluation disclosed a collagen fibrils orientation very close to the structure of a normal anterior cruciate ligament (ACL) where the collagen fibres are multidirectional [Strocchi et al. in J Anat 180(3):515-519, 1992]. On the other hand we found an unimodal distribution of collagen fibrils in the reconstructed ACL. This suggests that even at long-term follow-up stress exerts a variable influence. The multidirectional arrangement of collagen fibres resembles a normal ACL, but the unimodal distribution of fibrils is quite different from those seen in normal tendon and ligaments which tend to have a bimodal peak [Decker et al. in J Submicrosc Cytol Pathol 23:9-21, 1991; Strocchi et al. in J Anat 180(3):515-519, 1992]. Studies based on biopsy suffer from the potential weakness that the specimen may not have been representative of the entire prosthesis. Further long-term studies, possibly with the entire prosthesis and not only a biopsy, would highlight the behaviour and remodelling of this artificial ligament in greater detail and could be important for the development of future generations of artificial ligaments or tissue engineering ACL reconstruction.

Displayed correlation between gene expression profiles and submicroscopic alterations in response to cetuximab, gefitinib and EGF in human colon cancer cell lines.

BACKGROUND: EGFR is frequently overexpressed in colon cancer. We characterized HT-29 and Caco-2, human colon cancer cell lines, untreated and treated with cetuximab or gefitinib alone and in combination with EGF. METHODS: Cell growth was determined using a variation on the MTT assay. Cell-cycle analysis was conducted by flow cytometry. Immunohistochemistry was performed to evaluate EGFR expression and scanning electron microscopy (SEM) evidenced the ultrastructural morphology. Gene expression profiling was performed using hybridization of the microarray Ocimum Pan Human 40 K array A. RESULTS: Caco-2 and HT-29 were respectively 66.25 and 59.24 % in G0/G1. They maintained this level of cell cycle distribution after treatment, suggesting a predominantly differentiated state. Treatment of Caco-2 with EGF or the two EGFR inhibitors produced a significant reduction in their viability. SEM clearly showed morphological cellular transformations in the direction of cellular death in both cell lines treated with EGFR inhibitors. HT-29 and Caco-2 displayed an important reduction of the microvilli (which also lose their erect position in Caco-2), possibly invalidating microvilli absorption function. HT-29 treated with cetuximab lost their boundary contacts and showed filipodi; when treated with gefitinib, they showed some vesicles: generally membrane reshaping is evident. Both cell lines showed a similar behavior in terms of on/off switched genes upon treatment with cetuximab. The gefitinib global gene expression pattern was different for the 2 cell lines; gefitinib treatment induced more changes, but directly correlated with EGF treatment. In cetuximab or gefitinib plus EGF treatments there was possible summation of the morphological effects: cells seemed more weakly affected by the transformation towards apoptosis. The genes appeared to be less stimulated than for single drug cases. CONCLUSION: This is the first study to have systematically investigated the effect of cetuximab or gefitinib, alone and in combination with EGF, on human colon cancer cell lines. The EGFR inhibitors have a weaker effect in the presence of EGF that binds EGFR. Cetuximab treatment showed an expression pattern that inversely correlates with EGF treatment. We found interesting cyto-morphological features closely relating to gene expression profile. Both drugs have an effect on differentiation towards cellular death.

Polylactic acid fibre-reinforced polycaprolactone scaffolds for bone tissue engineering.

The employment of composite scaffolds with a well-organized architecture and multi-scale porosity certainly represents a valuable approach for achieving a tissue engineered construct to reproduce the middle and long-term behaviour of hierarchically complex tissues such as spongy bone. In this paper, fibre-reinforced composites scaffold for bone tissue engineering applications is described. These are composed of poly-L-lactide acid (PLLA) fibres embedded in a porous poly(epsilon-caprolactone) matrix, and were obtained by synergistic use of phase inversion/particulate leaching technique and filament winding technology. Porosity degree as high as 79.7% was achieved, the bimodal pore size distribution showing peaks at ca 10 and 200 microm diameter, respectively, accounting for 53.7% and 46.3% of the total porosity. In vitro degradation was carried out in PBS and SBF without significant degradation of the scaffold after 35 days, while in NaOH solution, a linear increase of weight lost was observed with preferential degradation of PLLA component. Subsequently, marrow stromal cells (MSC) and human osteoblasts (HOB) reached a plateau at 3 weeks, while at 5 weeks the number of cells was almost the same. Human marrow stromal cell and trabecular osteoblasts rapidly proliferate on the scaffold up to 3 weeks, promoting an oriented migration of bone cells along the fibre arrangement. Moreover, the role of seeded HOB and MSC on composite degradation mechanism was assessed by demonstrating a more relevant contribution to PLLA degradation of MSC when compared to HOB. The novel PCL/PLLA composite scaffolds thus showed promise whenever tuneable porosity, controlled degradability and guided cell-material interaction are simultaneously requested.

The role of hydroxyapatite as solid signal on performance of PCL porous scaffolds for bone tissue regeneration.

Highly porous composites made up of biodegradable poly-epsilon-caprolactone (PCL) and stoichiometric hydroxyapatite (HA) particles have been developed as substrate for bone-tissue regeneration. The processing technique consists of phase inversion and particulate (salt crystals) leaching. Three different HA contents (13, 20 and 26 vol %) in PCL-based composite were considered in this study. Pore microstructure with fully interconnected network and pore sizes ranging around a few hundred of mum (macroporosity) was obtained as a result of salt particles removal by leaching process. Several microns (microporosity) porosity was also created through phase inversion of polymer solution. Total porosity up to 95% was achieved. Human marrow stromal cells (MSC) were seeded onto porous PCL-based composites for 1-5 weeks and cultured in osteogenic medium. MSC were able to adhere and grow on PCL-based substrates with a plateau at 3-4 weeks. However, the small effect of bioactive signals on the biological response evaluated in MSC cell culture suggests a prior role of topography on the biological response. Importantly, the presence of HA as a bioactive solid signal determines an increase of mechanical properties. On the overall, the results indicated that porous PCL-based composites are potential candidate for bone substitution with beneficial influence on structural characteristics by solid signal addition.

Decorin transfection induces proteomic and phenotypic modulation in breast cancer cells 8701-BC.

Decorin is a prototype member of the small leucine-rich proteoglycan family widely distributed in the extracellular matrices of many connective tissues, where it has been shown to play multiple important roles in the matrix assembly process, as well as in some cellular activities. A major interest for decorin function concerns its role in tumorigenesis, as growth-inhibitor of different neoplastic cells, and potential antimetastatic agent. The aim of our research was to investigate wide-ranged effects of transgenic decorin on breast cancer cells. To this purpose we utilized the well-characterized 8701-BC cell line, isolated from a ductal infiltrating carcinoma of the breast, and two derived decorin-transfected clones, respectively, synthesizing full decorin proteoglycan or its protein core. The responses to the ectopic decorin production were examined by studying morphological changes, cell proliferation rates, and proteome modulation. The results revealed new important antioncogenic potentialities, likely exerted by decorin through a variety of distinct biochemical pathways. Major effects included the downregulation of several potential breast cancer biomarkers, the reduction of membrane ruffling, and the increase of cell-cell adhesiveness. These results disclose original aspects related to the reversion of malignant traits of a prototype of breast cancer cells induced by decorin. They also raise additional interest for the postulated clinical application of decorin.

Influence of different implant surfaces on peri-implant osteogenesis: histomorphometric analysis in sheep.

BACKGROUND: The present study investigated peri-implant osteogenesis and implant biologic fixation in different zirconia sandblasted endosseous titanium surfaces (SLA-60 and SLA-120) and a turned titanium surface (T) 2 and 4 weeks after surgery. METHODS: Seventy-two implant screws were implanted in tibia of six sheep. Histologic sections of implants (2 and 4 weeks after surgery) were analyzed with light microscopy for histomorphometric analysis of bone-to-implant contact (BIC), bone ingrowth (BI), and bone surface (BS/BV). Histologic blocks were used to perform bone microhardness studies next to the implants. Some implants were also observed with scanning electron microscopy (SEM) and transmission electron microscopy (TEM). RESULTS: In general, the highest values of BIC, BI, BS/BV, and Vickers hardness number (HV) were measured in SLA-60 samples, followed by SLA-120 and T implants. Two weeks after surgery, all the implants appeared biologically fixed by a newly formed woven bone arranged in thin bone trabeculae and filling the gap between implant and host bone. Four weeks after implantation, the thickness of the woven bone trabeculae had increased, especially around the SLA-60 and SLA-120 implants by a gradual deposition of parallel-fiber bone. CONCLUSIONS: Our results suggest that, in the early period of peri-implant healing, the implant surface morphology that seemed to influence the increase of peri-implant osteogenesis, bone turnover, and peri-implant bone maturation was SLA-60. We suggest that this surface, characterized by moderately deep titanium cavities very similar to the osteocyte lacunae, could act as a microscopic scaffold for mesenchymal and/or osteoblast-like cells adhesion.

Comparison of human mandibular osteoblasts grown on two commercially available titanium implant surfaces.

BACKGROUND: Surface characteristics play a major role in determining tissue response to implants and therefore their clinical outcome. The aim of the present study was to compare two commercially available titanium surfaces: plasma sprayed (TPS) and sand-blasted, acid-etched surface (SLA). METHODS: The surfaces were characterized by roughness testing, scanning electronic microscopy (SEM), Raman spectroscopy, and protein adsorption to determine their microtopographic and chemical properties. The effect of the surfaces on human mandibular osteoblasts was then studied in terms of cell morphology, adhesion, proliferation, and differentiation. Human osteoblasts from the mandible were cultured on these two surfaces and evaluated at 3, 6, 24, and 48 hours to determine cell attachment and morphology. Growth and differentiation kinetics were subsequently investigated by evaluating cell growth, alkaline phosphatase activity, osteocalcin and osteoprotegerin production at 7, 14, and 21 days. RESULTS: Although roughness was quite similar, the two surfaces presented strong differences in their topography, and cell morphology varied as a consequence. Osteoblasts on SLA appeared more elongated and spindle shaped than those on TPS, and their adhesion at 3 and 6 hours was weaker, but reached that of cells on TPS at hour 24. Cell proliferation was greater on SLA surfaces but differentiation parameters; i.e., alkaline phosphatase and osteocalcin, provided better results on TPS surfaces. Osteoprotegerin production was enhanced on TPS surfaces at days 14 and 21. CONCLUSION: Although cells grown on both surfaces exhibited good adhesion capabilities, a well-differentiated osteoblastic phenotype, and maintained a clear proliferation potential, our study suggests that plasma-sprayed treatment offers a better performance than SLA by creating, at least in the early phases, better conditions for tissue healing.

Different titanium surface treatment influences human mandibular osteoblast response.

BACKGROUND: Six titanium disks with six different surface treatments were examined: SS: smooth (polished) surface; TPS: plasma spray; C100: sand blasting by aluminum oxide (Al2O3) diameter 100 microm and acid etching; C150: sand blasting by Al2O3 diameter 150 microm and acid etching; B60: sand blasting by zirconium oxide (ZrO2) diameter 60 microm and acid etching; and B120: sand blasting by ZrO2 diameter 120 microm and acid etching. METHODS: The surface characteristics were determined by scanning electron microscopy (SEM) observation and a roughness tester. Raman spectroscopy was used to determine the presence of residual substances on the samples. Cells were seeded onto the disk and after 24 hours, 6 days, and 12 days were observed under SEM and growth curves generated with a cell counter. Some samples were used to determine alkaline phosphatase activity (ALP), using a colorimetric assay. RESULTS: SEM observation revealed drastic differences in surface microtopography, with a higher cell density on sand-blasted and acid-etched (SLA) samples than SS and TPS, and more regularly aligned cells on B60 and B120 surfaces than on the others. The growth curves showed a greater adhesion of cells on the etched/blasted surfaces compared to the SS and TPS surfaces. The number of cells increased on all the SLA samples, especially B60, throughout the experiment. At the same time, there was considerable ALP activity on the B60 sample, while it remained at extremely low levels on SS and TPS surfaces. Raman analyses revealed Al2O3 debris on C100 and C150, partly explaining the poorer performances of these two surface treatments, since this substance was shown to be toxic for cultured osteoblasts. CONCLUSIONS: Surface treatments influence the growth and the metabolic activity of cultured osteoblasts, and B60 seems to be the most favorable surface inducing a more pronounced proliferation of cells together with a high differentiation degree.

Osteogenesis and morphology of the peri-implant bone facing dental implants.

This study investigated the influence of different implant surfaces on peri-implant osteogenesis and implant face morphology of peri-implant tissues during the early (2 weeks) and complete healing period (3 months). Thirty endosseous titanium implants (conic screws) with differently treated surfaces (smooth titanium = SS, titanium plasma sprayed = TPS, sand-blasted zirconium oxide = Zr-SLA) were implanted in femur and tibiae diaphyses of two mongrel sheep. Histological sections of the implants and surrounding tissues obtained by sawing and grinding techniques were observed under light microscopy (LM). The peri-implant tissues of other samples were mechanically detached from the corresponding implants to be processed for SEM observation. Two weeks after implantation, we observed osteogenesis (new bone trabeculae) around all implant surfaces only where a gap was present at the host bone-metal interface. No evident bone deposition was detectable where threads of the screws were in direct contact with the compact host bone. Distance osteogenesis predominated in SS implants, while around rough surfaces (TPS and Zr-SLA), both distance and contact osteogenesis were present. At SEM analysis 2 weeks after implantation, the implant face of SS peri-implant tissue showed few, thin, newly formed, bone trabeculae immersed in large, loose, marrow tissue with blood vessels. Around the TPS screws, the implant face of the peri-implant tissue was rather irregular because of the rougher metal surface. Zr-SLA screws showed more numerous, newly formed bone trabeculae crossing marrow spaces and also needle-like crystals in bone nodules indicating an active mineralising process. After 3 months, all the screws appeared osseointegrated, being almost completely covered by a compact, mature, newly formed bone. However, some marrow spaces rich in blood vessels and undifferentiated cells were in contact with the metal surface. By SEM analysis, the implant face of the peri-implant tissue showed different results. Around the SS screws, the compact bone with areas of different mineralisation rate appeared very smooth, while around the rougher TPS screws, the bone still showed an irregular surface corresponding to the implant macro/microroughness. Around the Zr-SLA screws, a more regular implant-bone surface and sparse, calcified marrow spaces were detectable. Results from this research suggest that 2 weeks after implantation, trabecular bone represents the calcified healing tissue, which supports the early biological fixation of the implants. The peri-implant marrow spaces, rich in undifferentiated cells and blood vasculature, observed both 2 weeks and 3 months after surgery, favour the biological turnover of both early and mature peri-implant bone. The implant surface morphology strongly influences the rate and the modality of peri-implant osteogenesis, as do the morphology and arrangement of the implant face in peri-implant bone both during early healing (after 2 weeks) and when the implant is just osseointegrated; rough surfaces, and in particular Zr-SLA, seem to better favour bone deposition on the metal surface.