Evaluation of the Effects of Low-Level Laser Therapy on Diabetic Bone Healing.

The aim of the present study was to evaluate the effects of low-level laser therapy (LLLT) and biphasic alloplastic bone graft material on diabetic bone healing. Induction of diabetes was performed in 14 male Sprague-Dawley rats by intraperitoneal injection of a 50 mg/kg dose of streptozotocin. Two bilaterally symmetrical non-critical-sized bone defects were created in the parietal bones in each rat. Right defects were filled with biphasic alloplastic bone graft. Rats were randomly divided into 2 groups, with 1 group receiving 10 sessions of LLLT (GaAlAs, 78.5 J/cm, 100mW, 0.028 cm beam). The LLLT was started immediately after surgery and once every 3 days during postoperative period. At the end of treatment period, new bone formation and osteoblast density were determined using histomorphometry. Empty (control), graft-filled, LLLT-treated and both graft-filled and LLLT-treated bone defects were compared. New bone formation was higher in the graft treatment samples compared with the control (P = 0.009) and laser samples (P = 0.029). In addition, graft-laser combination treatment samples revealed higher bone formation than control (P = 0.008) and laser (P = 0.026) samples. Osteoblast density was significantly higher in the laser treatment (P <0.001), graft treatment (P = 0.001) and graft-laser combination treatment (P <0.001) samples than control samples. In addition, significantly higher osteoblast density was observed in the graft-laser combination treatment samples compared to the graft treatment samples (P = 0.005). The LLLT was effective to stimulate osteoblastogenesis but failed to increase bone formation. Graft augmentation for treatment of bone defects seems essential for proper bone healing in diabetes, regeneration may be supported by the LLLT to enhance osteoblastogenesis.

Evaluating the bioactivity of a hydroxyapatite-incorporated polyetheretherketone biocomposite.

BACKGROUND: Polyetheretherketone (PEEK) exhibits stable chemical properties, excellent biocompatibility, and rational mechanical properties that are similar to those of human cortical bone, but the lack of bioactivity impedes its clinical application. METHODS: In this study, hydroxyapatite (HA) was incorporated into PEEK to fabricate HA/PEEK biocomposite using a compounding and injection-molding technique. The tensile properties of the prepared HA/PEEK composites (HA content from 0 to 40 wt%) were tested to choose an optimal HA content. To evaluate the bioactivity of the composite, the cell attachment, proliferation, spreading and alkaline phosphatase (ALP) activity of MC3T3-E1 cells, and apatite formation after immersion in simulated body fluid (SBF), and osseointegration in a rabbit cranial defect model were investigated. The results were compared to those from ultra-high molecular weight polyethylene (UHMWPE) and pure PEEK. RESULTS: By evaluating the tensile properties and elastic moduli of PEEK composite samples/PEEK composites with different HA contents, the 30 wt% HA/PEEK composite was chosen for use in the subsequent tests. The results of the cell tests demonstrated that PEEK composite samples/PEEK composite exhibited better cell attachment, proliferation, spreading, and higher ALP activity than those of UHMWPE and pure PEEK. Apatite islands formed on the HA/PEEK composite after immersion in SBF for 7 days and grew continuously with longer time periods. Animal tests indicated that bone contact and new bone formation around the HA/PEEK composite were more obvious than those around UHMWPE and pure PEEK. CONCLUSIONS: The HA/PEEK biocomposite created by a compounding and injection-molding technique exhibited enhanced osteogenesis and could be used as a candidate of orthopedic implants.

Violence in the Early Bronze Age. Diagnosis of skull lesions using anthropological, taphonomic and scanning electron microscopy techniques.

In this paper we present the study of a skull belonging to a young male from the Italian Bronze Age showing three perimortem injuries on the frontal and parietal bones; the peculiarity of the frontal injury is represented by its singular shape, which may be indicative of the weapon that caused the lesion. The aim of the present study is to examine the traumatic evidence in relation to possible etiological factors, in order to attempt to establish if the lesion occurred peri or post-mortem, and to evaluate if these traumatic injuries could be interpreted as an evidence of interpersonal violence, by combining anthropological, taphonomic and ESEM investigations. The combination of multidisciplinary methods of study can provide important new insights into inter-personal violence.

Adjunctive hyperbaric oxygen therapy for irradiated rat calvarial defects

PURPOSE: The aim of this study was to conduct a histologic evaluation of irradiated calvarial defects in rats 4 weeks after applying fibroblast growth factor-2 (FGF-2) with hyaluronan or biphasic calcium phosphate (BCP) block in the presence or absence of adjunctive hyperbaric oxygen (HBO) therapy. METHODS: Twenty rats were divided into HBO and non-HBO (NHBO) groups, each of which was divided into FGF-2 and BCP-block subgroups according to the grafted material. Localized radiation with a single 12-Gy dose was applied to the calvaria of rats to simulate radiotherapy. Four weeks after applying this radiation, 2 symmetrical circular defects with a diameter of 6 mm were created in the parietal bones of each animal. The right-side defect was filled with the materials mentioned above and the left-side defect was not filled (as a control). All defects were covered with a resorbable barrier membrane. During 4 weeks of healing, 1 hour of HBO therapy was applied to the rats in the HBO groups 5 times a week. The rats were then killed, and the calvarial specimens were harvested for radiographic and histologic analyses. RESULTS: New bone formation was greatest in the FGF-2 subgroup, and improvement was not found in the BCP subgroup. HBO seemed to have a minimal effect on new bone formation. There was tendency for more angiogenesis in the HBO groups than the NHBO groups, but the group with HBO and FGF-2 did not show significantly better outcomes than the HBO-only group or the NHBO group with FGF-2. CONCLUSIONS: HBO exerted beneficial effects on angiogenesis in calvarial defects of irradiated rats over a 4-week healing period, but it appeared to have minimal effects on bone regeneration. FGF-2 seemed to enhance new bone formation and angiogenesis, but its efficacy appeared to be reduced when HBO was applied.

Eight-week healing of grafted calvarial bone defects with hyperbaric oxygen therapy in rats

PURPOSE: The purpose of this study was to evaluate the synergistic effect of adjunctive hyperbaric oxygen (HBO) therapy on new bone formation and angiogenesis after 8 weeks of healing. METHODS: Sprague-Dawley rats (n=28) were split into 2 groups according to the application of adjunctive HBO therapy: a group that received HBO therapy (HBO group [n=14]) and another group that did not receive HBO therapy (NHBO group [n=14]). Each group was divided into 2 subgroups according to the type of bone graft material: a biphasic calcium phosphate (BCP) subgroup and an Escherichia coli-derived recombinant human bone morphogenetic protein-2-/epigallocatechin-3-gallate-coated BCP (mBCP) subgroup. Two identical circular defects with a 6-mm diameter were made in the right and left parietal bones of each rat. One defect was grafted with bone graft material (BCP or mBCP). The other defect was not grafted. The HBO group received 2 weeks of adjunctive HBO therapy (1 hour, 5 times a week). The rats were euthanized 8 weeks after surgery. The specimens were prepared for histologic analysis. RESULTS: New bone (%) was higher in the NHBO-mBCP group than in the NHBO-BCP and control groups (P<0.05). Blood vessel count (%) and vascular endothelial growth factor staining (%) were higher in the HBO-mBCP group than in the NHBO-mBCP group (P<0.05). CONCLUSIONS: HBO therapy did not have a positive influence on bone formation irrespective of the type of bone graft material applied after 8 weeks of healing. HBO therapy had a positive effect on angiogenic activity.

Photobiomodulation of mesenchymal stem cells encapsulated in an injectable rhBMP4-loaded hydrogel directs hard tissue bioengineering.

Photobiomodulation (PBM) therapy displays relevant properties for tissue healing and regeneration, which may be of interest for the tissue engineering field. Here, we show that PBM is able to improve cell survival and to interact with recombinant human Bone Morphogenetic Protein 4 (rhBMP4) to direct and accelerate odonto/osteogenic differentiation of dental derived mesenchymal stem cells (MSCs). MSCs were encapsulated in an injectable and thermo-responsive cell carrier (Pluronic® F-127) loaded with rhBMP4 and then photoactivated. PBM improved MSCs self-renewal and survival upon encapsulation in the Pluronic® F-127. In the presence of rhBMP4, cell odonto/osteogenic differentiation was premature and markedly improved in the photoactivated MSCs. An in vivo calvarial critical sized defect model demonstrated significant increase in bone formation after PBM treatment. Finally, a balance in the reactive oxygen species levels may be related to the favorable results of PBM and rhBMP4 association. PBM may act in synergism with rhBMP4 and is a promise candidate to direct and accelerate hard tissue bioengineering.

Four-week histologic evaluation of grafted calvarial defects with adjunctive hyperbaric oxygen therapy in rats

PURPOSE: The aim of this study was to characterize the healing in the grafted calvarial defects of rats after adjunctive hyperbaric oxygen therapy. METHODS: Twenty-eight male Sprague-Dawley rats (body weight, 250–300 g) were randomly divided into two treatment groups: with hyperbaric oxygen therapy (HBO; n=14) and without HBO (NHBO; n=14). Each group was further subdivided according to the bone substitute applied: biphasic calcium phosphate (BCP; n=7) and surface-modified BCP (mBCP; n=7). The mBCP comprised BCP coated with Escherichia-coli-derived recombinant human bone morphogenetic protein-2 (ErhBMP-2) and epigallocatechin-3-gallate (EGCG). Two symmetrical circular defects (6-mm diameter) were created in the right and left parietal bones of each animal. One defect was assigned as a control defect and received no bone substitute, while the other defect was filled with either BCP or mBCP. The animals were allowed to heal for 4 weeks, during which those in the HBO group underwent 5 sessions of HBO. At 4 weeks, the animals were sacrificed, and the defects were harvested for histologic and histomorphometric analysis. RESULTS: Well-maintained space was found in the grafted groups. Woven bone connected to and away from the defect margin was formed. More angiogenesis was found with HBO and EGCG/BMP-2 (P<0.05). None of the defects achieved complete defect closure. Increased new bone formation with HBO or EGCG/BMP-2 was evident in histologic evaluation, but it did not reach statistical significance in histometric analysis. A synergic effect between HBO and EGCG/BMP-2 was not found. CONCLUSIONS: Within the limitations of this study, the present findings indicate that adjunctive HBO and EGCG/BMP-2 could be beneficial for new bone formation in rat calvarial defects.

Histomorphometric and microtomographic evaluation of the effects of hyperbaric oxygen and systemic ozone, used alone and in combination, on calvarial defect healing in rats.

PURPOSE: The aim of this study was to compare the efficacy of hyperbaric oxygen and systemic ozone, used separately and in combination, on the healing of bone defects. MATERIAL AND METHODS: Sixty male Wistar rats were divided into 4 groups (n = 15) according to treatment (control, hyperbaric oxygen [HBO], ozone [O], and HBO plus O [HBO-O]) and divided further into 3 subgroups according to day of sacrifice (postsurgical days 5, 15, and 30). Surgery was performed under general anesthesia to create a critical-size bone defect (5 mm in diameter) in the cranium. After sacrifice, microtomographic images of all samples were recorded, and histomorphometric analysis was performed. RESULTS: Histologic and radiologic measurements showed that the values of all experimental groups were higher than those of the control group. Histologic scores for all experimental groups were statistically higher than those for the control group day 30 (O, P = .045; HBO, P = .049; HBO-O, P = .042). Histologic scores also were statistically higher for the HBO group on day 5 (P = .045) and day 15 (P = .009) compared with the control group. Microtomographic scores were higher for the experimental groups than for the control group, with statistically significant differences for group O on day 5 (P = .033) and day 30 (P = .0045) and for group HBO on day 15 (P = .005). Histologic and radiologic analyses showed positive correlations. CONCLUSION: Within the limitations of this study, the use of hyperbaric oxygen and ozone, separately and in combination, were shown to be effective in increasing bone healing. Combined usage was no more effective in stimulating bone healing than separate usage.

The effects of tissue-non-specific alkaline phosphatase gene therapy on craniosynostosis and craniofacial morphology in the FGFR2C342Y/+ mouse model of Crouzon craniosynostosis.

OBJECTIVES: Craniosynostosis, the premature fusion of cranial bones, has traditionally been described as a disease of increased bone mineralization. However, multiple mouse models of craniosynostosis display craniosynostosis simultaneously with diminished cranial bone volume and/or density. We propose an alternative hypothesis that craniosynostosis results from abnormal tissue mineralization through the downregulation of tissue-non-specific alkaline phosphatase (TNAP) enzyme downstream of activating mutations in FGFRs. MATERIAL AND METHODS: Neonatal Crouzon (FGFRC342Y/+) and wild-type (FGFR+/+) mice were injected with lentivirus to deliver a recombinant form of TNAP. Mice were sacrificed at 4 weeks postnatal. Serum was collected to test for alkaline phosphatase (AP), phosphorus, and calcium levels. Craniofacial bone fusion and morphology were assessed by micro-computed tomography. RESULTS: Injection with the TNAP lentivirus significantly increased serum AP levels (increased serum AP levels are indicative of efficient transduction and production of the recombinant protein), but results were variable and dependent upon viral lot and the litter of mice injected. Morphological analysis revealed craniofacial form differences for inferior surface (p=0.023) and cranial height (p=0.014) regions between TNAP lentivirus-injected and vehicle-injected Crouzon mice. With each unit increase in AP level, the odds of lambdoid suture fusion decreased by 84.2% and these results came close to statistical significance (p=0.068). CONCLUSION: These results suggest that TNAP deficiency may mediate FGFR2-associated craniosynostosis. Future studies should incorporate injection of recombinant TNAP protein, to avoid potential side effects and variable efficacy of lentiviral gene delivery.

Clumps of a mesenchymal stromal cell/extracellular matrix complex can be a novel tissue engineering therapy for bone regeneration.

BACKGROUND AIMS: The transplantation of mesenchymal stromal cells (MSCs) to damaged tissue has attracted attention in scientific and medical fields as an effective regenerative therapy. Nevertheless, additional studies are required to develop an MSC transplant method for bone regeneration because the use of an artificial scaffold restricts the number of transplanted cells and their function. Furthermore, regulating the degree of cell differentiation in vitro is desirable for a more effective regenerative therapy. To address these unresolved issues, with the use of a self-produced extracellular matrix (ECM), we developed clumps of an MSC/ECM complex (C-MSCs). METHODS: MSCs isolated from rat femur were cultured in growth medium supplemented with 50 µg/mL of ascorbic acid for 7 days. To obtain C-MSCs, confluent cells were scratched with the use of a micropipette tip to roll up the cellular sheet, which consisted of ECM produced by the MSCs. The biological properties of C-MSCs were assessed in vitro and their bone regenerative activity was tested by use of a rat calvarial defect model. RESULTS: Immunofluorescent confocal microscopic analysis revealed that type I collagen formed C-MSCs. Osteopontin messenger RNA expression and amount of calcium content were higher in C-MSCs cultured in osteo-inductive medium than those of untreated C-MSCs. The transplantation of osteogenic-differentiated C-MSCs led to rapid bone regeneration in the rat calvarial defect model. CONCLUSIONS: These results suggest that the use of C-MSCs refined by self-produced ECM, which contain no artificial scaffold and can be processed in vitro, may represent a novel tissue engineering therapy.

Calcification during bone healing in a standardised rat calvarial defect assessed by micro-CT and SEM-EDX.

OBJECTIVE: The study was designed to investigate the process of calcification during bone healing in a standardized rat calvarial bone defect model, measured by bone mineral density and the concentrations and distributions of calcium, phosphorus and carbon in the bone matrix. MATERIALS AND METHODS: A standard defect was made on the parietal bone of 12-week-old rats under anaesthesia. The rats were fixed in weeks 1, 2, 4 and 8,and the calvaria were resected and examined with microcomputed tomography, then frozen and sectioned for histology and analysed with energy-dispersive X-ray spectroscopy (EDX). Parietal bone of 12-week-old control rats was processed similarly. RESULTS: The mineral density of healing bone increased with time. The healing bone became thicker and denser with time in histology. The distributions of Ca and P expanded over the bone matrix, whereas that of C became localised and complemented that of C and P. The Ca/P concentration ratio increased, whereas the C/Ca and C/P ratios decreased in the healing bone matrix. CONCLUSION: Healing bone is immaturely calcified initially and proceeds calcification gradually, that is, as the bone volume increases, mineral increases in density and matures in quality, while organic components decrease.

A re-examination of the human fossil specimen from Backi Petrovac (Serbia).

A fragmented human calotte was discovered during the early 1950s near Backi Petrovac (Serbia), in association with Palaeolithic stone tools. After its initial publication, the fossil specimen remained largely unknown outside of the Serbian academe and no detailed comparative study has ever been carried out. Since the whereabouts of the fossil itself are currently unknown, and given its potential significance for the Pleistocene human evolution, we re-examine the data published by Zivanovic (1966, 1975). Using the original measurements, mostly taken on the frontal bone, and a wide comparative sample of 68 fossil specimens, the fossil was compared and analyzed by statistical multivariate methods. We also conducted a visual examination of the morphology based on the available photographic material. Our analysis reveals phenetic similarity with Middle Pleistocene archaic Homo from Africa and anatomically modern Homo sapiens. However, the absence of primitive cranial traits in Backi Petrovac indicates a clear modern Homo sapiens designation. Although lost at the moment, there is a chance for the re-discovery of the fossil in the years to come. This would give us an opportunity to acquire absolute dates and to study the specimen in a more detailed manner.

Fatal cranial injury in an individual from Messina (Sicily) during the times of the Roman Empire.

Forensic and archaeological examinations of human skeletons can provide us with evidence of violence. In this paper, we present the patterns of two cranial lesions found on an adult male (T173) buried in a grave in the necropolis 'Isolato 96', Messina, Sicily, dating back to the Roman Empire (1st century BC - 1st century AD). The skull reveals two perimortem traumatic lesions, one produced by a sharp object on the right parietal bone and the other one on the left parietal bone, presumably the result of a fall. The interpretation of fracture patterns found in this cranium are an illustration of how forensic approaches can be applied with great benefit to archaeological specimens.

Effects of laser and ozone therapies on bone healing in the calvarial defects.

This study aims to analyze the effect of the low-level laser therapy (LLLT) and ozone therapy on the bone healing of critical size defect (CSD) in rat calvaria. A total of 30 Wistar male rats were used. A 5-mm-diameter trephine bur was used to create CSD on the right side of the parietal bone of each rat calvarium. Once the bone was excised, a synthetic biphasic calcium phosphate graft material was implanted to all the bone defect sites. The animals were randomly divided into 3 groups as follows: the control group (n = 10), which received no LLLT or ozone therapy; the LLLT group (n = 10), which received only LLLT (120 seconds, 3 times a week for 2 weeks); and the ozone therapy group (n = 10) (120 seconds, 3 times a week for 2 weeks). After 1 month, all the rats were killed, and the sections were examined to evaluate the presence of inflammatory infiltrate, connective tissue, and new bone formation areas. Histomorphometric analyses showed that in the LLLT and ozone groups, the new bone areas were significantly higher than in the control group (P < 0.05). In the LLLT group, higher new bone areas were found than in the ozone group (P < 0.05). This study demonstrated that both ozone and laser therapies had a positive effect on bone formation in rat calvarial defect, compared with the control group; however, ozone therapy was more effective than LLLT (808 nm; 0.1 W; 4 J/cm(2); 0.028 cm(2), continuous wave mode).

Effects of cigarette smoke inhalation and coffee consumption on bone formation and osseous integration of hydroxyapatite implant.

The present study aims to assess the effects of cigarette smoke inhalation and/or coffee consumption on bone formation and osseous integration of a dense hydroxyapatite (DHA) implant in rats. For this study, 20 male rats were divided into four groups (n = 5): CT (control) group, CE (coffee) group, CI (cigarette) group and CC (coffee + cigarette) group. During 16 weeks, animals in the CI group were exposed to cigarette smoke inhalation equivalent to 6 cigarettes per day; specimens in the CE group drank coffee as liquid diet; and rats in the CC group were submitted to both substances. In the 6th week a 5 mm slit in the parietal bone and a 4 mm slit in the tibia were performed on the left side: the former was left open while the latter received a DHA implant. As soon as surgeries were finished, the animals returned to their original protocols and after 10 weeks of exposure they were euthanised (ethically sacrificed) and the mentioned bones collected for histological processing. Data showed that exposure to cigarette smoke inhalation and coffee consumption did not interfere in weight gain and that solid and liquid diet consumption was satisfactory. Rats in the CC group showed a decrease in bone neoformation around the tibial DHA implant (31.8 ± 2.8) as well as in bone formation in the parietal slit (28.6 ± 2.2). On their own, cigarette smoke inhalation or coffee consumption also led to diminished bone neoformation around the implant and delayed the bone repair process in relation to the CT group. However, reduction in the bone repair process was accentuated with exposure to both cigarette smoke inhalation and coffee consumption in this study.

Hyperbaric oxygen stimulates vascularization and bone formation in rat calvarial defects.

Hyperbaric oxygen (HBO) therapy is used to treat or prevent tissue necrosis in patients undergoing irradiation. Many such patients require reconstructive surgery, but little is known of the effects of HBO on bone vascularization and regeneration. In this study, copolymer poly(l-lactide-co-1,5-dioxepan-2-one) (poly(LLA-co-DXO)) scaffolds were implanted into critical-sized calvarial defects in Wistar rats. The animals were randomly allotted to hyperbaric or normobaric oxygen groups. The treatment group received five sessions weekly for 90 min at increased atmospheric pressure, for up to 4 weeks. Samples were retrieved at weeks 2 and 8, i.e. after a total of 10 and 20 sessions, respectively. The samples were analyzed by real-time reverse transcriptase polymerase chain reaction (RT-PCR) and histology at week 2, and radiographically and histologically at week 8. At week 2, defects treated with HBO exhibited greater numbers of cells positive for the endothelial marker CD31, up-regulated gene expression of osteogenic markers, and down-regulated expression of pro-inflammatory cytokines. At week 8, radiographic examination revealed that calvarial defects subjected to HBO exhibited a higher percentage of radiopacities than normobaric controls, and histological examination disclosed enhanced bone healing. These results confirmed that HBO treatment was effective in stimulating vascularization and bone formation in rat calvarial defects.

Effects of cigarette smoke inhalation and coffee consumption on bone formation and osseous integration of hydroxyapatite implant / Os efeitos do cigarro e do consumo de café sobre a formação óssea e a integração óssea de implantes de hidroxiapatita

The present study aims to assess the effects of cigarette smoke inhalation and/or coffee consumption on bone formation and osseous integration of a dense hydroxyapatite (DHA) implant in rats. For this study, 20 male rats were divided into four groups (n = 5): CT (control) group, CE (coffee) group, CI (cigarette) group and CC (coffee + cigarette) group. During 16 weeks, animals in the CI group were exposed to cigarette smoke inhalation equivalent to 6 cigarettes per day; specimens in the CE group drank coffee as liquid diet; and rats in the CC group were submitted to both substances. In the 6th week a 5 mm slit in the parietal bone and a 4 mm slit in the tibia were performed on the left side: the former was left open while the latter received a DHA implant. As soon as surgeries were finished, the animals returned to their original protocols and after 10 weeks of exposure they were euthanised (ethically sacrificed) and the mentioned bones collected for histological processing. Data showed that exposure to cigarette smoke inhalation and coffee consumption did not interfere in weight gain and that solid and liquid diet consumption was satisfactory. Rats in the CC group showed a decrease in bone neoformation around the tibial DHA implant (31.8 ± 2.8) as well as in bone formation in the parietal slit (28.6 ± 2.2). On their own, cigarette smoke inhalation or coffee consumption also led to diminished bone neoformation around the implant and delayed the bone repair process in relation to the CT group. However, reduction in the bone repair process was accentuated with exposure to both cigarette smoke inhalation and coffee consumption in this study.

O presente estudo teve como objetivo avaliar os efeitos do tabagismo e do consumo de café, isolada ou concomitantemente, sobre a formação óssea e a osseointegração de implantes hidroxiapatita densa. Foram utilizados 20 ratos machos, divididos em quatro grupos (n = 5): grupo CT (controle); grupo CA (café); grupo CI (cigarro), e grupo CC (cigarro + café). Durante 16 semanas, os animais do grupo CI foram expostos à fumaça de seis cigarros/dia; os animais do grupo CA consumiram café como dieta líquida, e os animais do grupo CC, ambas as substâncias. Após seis semanas de exposição, uma falha óssea de 5 mm foi produzida no osso parietal esquerdo e de 4 mm, na tíbia esquerda dos animais. A falha do parietal foi mantida aberta, enquanto na tíbia corpos cerâmicos de hidroxiapatita densa (HAD) foram implantados em cavidade produzida cirurgicamente. Após as cirurgias, os animais retornaram aos protocolos experimentais e, ao término de dez semanas, foram eutanasiados, sendo as tíbias e os parietais coletados para processamento histológico. A exposição à fumaça do cigarro e o consumo de café não interferiram no ganho de peso dos animais, e os consumos de dieta líquida e sólida foram satisfatórios entre os grupos. Os animais do grupo CC apresentaram menor volume de osso neoformado ao redor do implante de HAD na tíbia (31,8 ± 2,8) e menor osteogênese na falha produzida no osso parietal (28,6 ± 2,2). O café e o cigarro consumidos isoladamente provocam a diminuição do volume de osso ao redor do implante e o atraso no processo de reparação óssea. Observou-se que o consumo de café associado à exposição à fumaça do cigarro reduziu de forma acentuada o processo de reparação óssea, no presente estudo.

Influence of low-level laser associated with osteogenic proteins recombinant human BMP-2 and Hevea brasiliensis on bone repair in Wistar rats.

This study analyzed the newly formed bone tissue after application of recombinant human BMP-2 (rhBMP-2) and P-1 (extracted from Hevea brasiliensis) proteins, 2 weeks after the creation of a critical bone defect in male Wistar rats treated or not with a low-intensity laser (GaAlAs 780 nm, 60 mW of power, and energy density dose of 30 J/cm(2)). The animals were divided into two major groups: (1) bone defect plus low-intensity laser treatment and (2) bone defect without laser irradiation. The following subgroups were also analyzed: (a) 5 µg of pure rhBMP-2; (b) 5 µg of pure P-1 fraction; (c) 5 µg of rhBMP-2/monoolein gel; (d) 5 µg of P-1 fraction/monoolein gel; (e) pure monoolein gel. Comparisons of the groups receiving laser treatment with those that did not receive laser irradiation show differences in the areas of new bone tissue. The group treated with 5 µg of rhBMP-2 and laser irradiation was not significantly different (P >0.05) than the nonirradiated group that received the same treatment. The irradiated, rhBMP-2/monoolein gel treatment group showed a lower area of bone formation than the nonirradiated, rhBMP-2/gel monoolein treatment group (P < 0.001). The area of new bone tissue in the other nonirradiated and irradiated groups was not significantly different (P > 0.05). Furthermore, the group that received the 5 µg of rhBMP-2 application showed the greatest bone formation. We conclude that the laser treatment did not interfere with the area of new bone tissue growth and that the greatest stimulus for bone formation involved application of the rhBMP-2 protein.

Application of low-level laser irradiation (LLLI) and rhBMP-2 in critical bone defect of ovariectomized rats: histomorphometric evaluation.

OBJECTIVES: The aim of this study was to evaluate the osteogenic potential of recombinant human bone morphogenetic protein-2 (rhBMP-2) and low-level laser irradiation (LLLI), isolated or combined in critical bone defects (5 mm) in parietal bone using ovariectomized female rats as an experimental animal model. MATERIALS AND METHODS: Forty-nine female Wistar rats, bilaterally ovariectomized (OVX), were divided into seven treatment groups of seven animals each: (I) laser in a single application, (II) 7 µg of pure rhBMP-2, (III) laser and 7 µg of pure rhBMP-2, (IV) 7 µg of rhBMP-2/monoolein gel, (V) laser and 7 µg of rhBMP-2/monoolein gel, (VI) laser and pure monoolein gel, and (VII) critical bone defect controls. The low-level laser source used was a gallium aluminum arsenide semiconductor diode laser device (λ = 780 nm, D = 120 J/cm(2)). RESULTS: Groups II and III presented higher levels of newly formed bone than all other groups with levels of 40.57% and 40.39%, respectively (p < 0.05). The levels of newly formed bone of groups I, IV, V, and VI were similar with levels of 29.67%, 25.75%, 27.75%, and 30.64%, respectively (p > 0.05). The area of new bone formation in group VII was 20.96%, which is significantly lower than groups I, II, III, and VI. CONCLUSIONS: It was concluded that pure rhBMP-2 and a single dose of laser application stimulated new bone formation, but the new bone formation area was significantly increased when only rhBMP-2 was used. Additionally, the laser application in combination with other treatments did not influence the bone formation area.

No influence of alimentary zinc on the healing of calvarial defects filled with osteopromotive substances in rats.

Zinc has been demonstrated to play an important role in bone metabolism and is required for normal growth. However, no studies have investigated the influence of zinc on calvarial bone healing in aged or adult rats. The aim of the study was to evaluate whether alimentary zinc supplementation and depletion affect bone healing of calvarial defects implanted with osteopromotive substances in adult rats. Two 5 mm full thickness critical size bone defects were trephined in the central part of each parietal bone of 60 six-month-old male Wistar rats. The bone defects were filled with demineralized bone matrix (DBM), autogenous bone chips, or were left as unfilled controls. The rats were divided into three groups of 20 rats each and received a semi-synthetic diet containing 20, 60, or 120 mg zinc/kg. After 4 months, the biomechanical integrity of the healing defects was evaluated by a punch out test and the healed defects were examined with histomorphometry. Statistical analysis of the data was carried out by two-way analysis of variance and Wilcoxon's non-parametric signed rank test. Biomechanical testing revealed that the maximum load was significantly higher in DBM-filled defects than in those filled with autogenous bone, and that the defects filled with autogenous bone were stronger than the unfilled controls. The biomechanical findings indicated that the alimentary zinc content did not influence the healing of calvarial defects. No significant difference in maximum load could be established between the three diet groups for any of the filling materials, whereas the highest zinc supplement resulted in an increase in the relative extension on mineralizing surfaces in the control group. Thus, healing of adult rat calvarial defects is not influenced by alimentary zinc supplementation or depletion. Defects filled with DBM were significantly stronger and exhibited significantly more new bone formation than defects filled with autogenous bone or unfilled controls.