Platelet-Rich Fibrin Progressive Protocol: Third Generation of Blood Concentrates.

PURPOSE: Platelet-rich fibrin (PRF) has been used in several fields of dentistry to improve tissue healing. However, PRF from glass tubes results in a limited number of small membranes, increasing clinical difficulty and work time. The aim of this study was to evaluate cell and platelet amounts and biomechanical strength of PRF-giant membranes produced from plastic tubes without additives. MATERIAL AND METHODS: The investigators designed an ex vivo study, to compare 3 different centrifugation protocols for obtaining PRF: 700 × g/12 minutes (leukocyte and PRF [L-PRF]), 350 × g/14 minutes (GM350), and 60-700 × g more than 15 minutes total (progressive PRF [PRO-PRF]). We collected blood samples from 5 volunteers aged 25-54 years, over 3 different time periods (triplicate and paired study). From each venipuncture, 4 mL of blood was collected in vacutainers with ethylenediamine tetraacetic acid (EDTA) and approximately 104 mL in 12 plastic tubes without additives, which were separated into 3 groups, as per the centrifugation protocols (n = 5): L-PRF, GM350, and PRO-PRF. The PRF from the tubes of the same protocol was aspirated and 9 mL were placed in polylactic acid (PLA) forms and 3 mL were placed in a glass receptacle. The membranes from PLA forms were tested for tensile strength and the membranes from glass receptacles were evaluated by histomorphometry, while platelets and leukocytes were counted for those in tubes with EDTA. Statistical analyses were performed using Shapiro-Wilk normality test and then a one-way repeated measures analysis followed by Tukey multiple comparisons test (α < 0.05). RESULTS: In tensile analyses, PRO-PRF (0.85 ± 0.23 N) showed a significantly higher maximum breaking strength than L-PRF (0.61 ± 0.26 N, P = .01) and GM350 (0.58 ± 0.23 N, P < .01). The histomorphometry revealed no significant statistical difference in cell counts between the groups (P = .52). Furthermore, there was no significant difference between the leukocyte (P = .25) and platelet counts (P = .59) in whole blood between the groups. CONCLUSION: The progressive protocol (PRO-PRF) enabled the production of PRF giant membranes with greater tensile strength and adequate cell distribution. Moreover, it allows biomaterial incorporation during production and enables clinical control of membrane thickness and size as per the surgical procedure.

Do blood concentrates influence inflammatory signs and symptoms after mandibular third molar surgery? A systematic review and network meta-analysis of randomized clinical trials.

OBJECTIVES: To investigate, through a network meta-analysis, the effectiveness of blood concentrates in reducing pain perception, trismus, and edema after mandibular third molar extraction. MATERIALS AND METHODS: An electronic search was performed in nine databases to locate randomized clinical trials comparing blood concentrate use after mandibular third molar extraction. Two authors selected and extracted the data independently. The individual risk of bias in the studies was assessed with the RoB v2.0 tool. A network meta-analysis compared postoperative pain and trismus scores after applying different blood concentrates, using the mean difference (MD) as an effect estimate. The GRADE approach assessed the certainty of evidence. RESULTS: Thirty-one randomized clinical trials were included in the review and 18 in the meta-analysis. Leukocyte- and platelet-rich fibrin (L-PRF) was the most used blood concentrate, followed by platelet-rich plasma (PRP). The network meta-analysis, depending on the analyzed period, evaluated up to 1240 surgeries. Among the analyzed blood concentrates, advanced platelet-rich fibrin (A-PRF) performed better among the analyzed blood concentrates, decreasing postoperative pain in 1, 2, 3, and 7 days and reducing trismus up to the inflammatory peak compared to blood clots. Only two studies had a low risk of bias. CONCLUSIONS: Based on very low certainty of evidence, using concentrates seemed efficient compared to blood clots in reducing pain and trismus after mandibular third molar surgeries. A-PRF decreased postoperative pain throughout the evaluated time and trismus during the acute inflammatory peak. CLINICAL RELEVANCE: A-PRF after mandibular third molar extractions performed better among the analyzed blood concentrates and seemed efficient in improving postoperative quality by decreasing inflammatory signs and symptoms.

Automatic Segmentation of Bone Canals in Histological Images.

The literature provides many works that focused on cell nuclei segmentation in histological images. However, automatic segmentation of bone canals is still a less explored field. In this sense, this paper presents a method for automatic segmentation approach to assist specialists in the analysis of the bone vascular network. We evaluated the method on an image set through sensitivity, specificity and accuracy metrics and the Dice coefficient. We compared the results with other automatic segmentation methods (neighborhood valley emphasis (NVE), valley emphasis (VE) and Otsu). Results show that our approach is proved to be more efficient than comparable methods and a feasible alternative to analyze the bone vascular network.

Effects of ionizing radiation on woven bone: influence on the osteocyte lacunar network, collagen maturation, and microarchitecture.

OBJECTIVES: Evaluate the effects of ionizing radiation on microarchitecture, the osteocyte lacunar network, and collagen maturity in a bone repair site. MATERIALS AND METHODS: Bone defects were created on tibias of 20 New Zealand rabbits. After 2 weeks, the animals were randomly divided into (n = 10) NoIr (nonirradiated group) and Ir (irradiated group). In the Ir, the animals received single-dose irradiation of 30 Gy on the tibia and were euthanized after 2 weeks. Bone microarchitecture parameters were analyzed by using micro-CT, and the osteocyte lacunar network, bone matrix, and collagen maturation by histomorphometric analysis. The data were analyzed using unpaired Student's t test (α = 0.05). RESULTS: Trabecular thickness in Ir was lower than that in NoIr (P = 0.028). No difference was found for bone volume fraction and bone area. Lacunae filled with osteocytes were more numerous (P < 0.0001) in NoIr (2.6 ± 0.6) than in Ir (1.97 ± 0.53). Empty lacunae were more prevalent (P < 0.003) in Ir (0.14 ± 0.10) than in NoIr (0.1 ± 0.1). The mean osteocyte lacunae size was higher (P < 0.01) in Ir (15.4 ± 4.41) than in NoIr (12.7 ± 3.7). Picrosirius red analysis showed more (P < 0.05) mature collagen in NoIr (29.0 ± 5.3) than in Ir (23.4 ± 4.5). Immature collagen quantification revealed no difference between groups. CONCLUSIONS: Ionizing radiation compromised bone formation and an impairment in bone repair in irradiated woven bone was observed. CLINICAL RELEVANCE: Before radiotherapy, patients usually need surgical intervention, which may be better performed, if clinicians understand the repair process in irradiated bone, using novel approaches for treating these individuals.

Impact of Childhood Obesity on Bone Metabolism.

Childhood obesity is a considerable worldwide health problem and a major risk factor for several chronic diseases. Fat rich diets result in altered serum levels of lipids, cytokines and hormonal factors, which influence skeletal acquisition and promote microstructural and mechanical behavior changes in bone, especially to bone quality and quantity. However, the possible longterm implications of high-fat diets in childhood are controversial. Despite not fully understood, multiple signaling pathways which support bone tissue homeostasis are altered under hyperlipidic conditions, including RANKL/RANK/OPG, PPAR-γ/Alox5/5-LO, leptin/IGF-I/AGE, ApoE/Lrp-1, Thy-1, IL-6, TNFα, calcium, vitamin D and K metabolism. Moreover, the expression of reactive oxygen species is also modified. Considering the importance of this subject, the aim of this review was to explore the mechanisms of bone formation affected by obesity during childhood during childhood.

Protective effects of a polyphenol-enriched fraction of the fruit peel of Annona crassiflora Mart. on acute and persistent inflammatory pain.

Different parts of Annona crassiflora Mart., a native species from Brazilian savanna, were traditionally used for the treatment of a wide variety of ailments including arthritis. Thus, this study aimed to investigate the possible antinociceptive and anti-inflammatory properties of a polyphenol-enriched fraction of the fruit peel of A. crassiflora, named here as EtOAc, in mice. Pro-inflammatory cytokines and nitric oxide (NO) production were evaluated in LPS-activated macrophages. Then, EtOAc fraction was administered by oral route in male C57BL/6/J mice, and the animals were submitted to glutamate-induced nociception and complete Freund's adjuvant (CFA)-induced monoarthritis tests to assess nociception (mechanical, spontaneous and cold pain) and inflammation (edema and neutrophil infiltration), and to the open-field and rotarod tests for motor performance analysis. EtOAc fraction inhibited the production of IL-6 and NO in the LPS-induced macrophages, and reduced spontaneous nociception induced by glutamate, without altering the animals' locomotor activity. In addition, the polyphenol-enriched fraction was able to revert the early and late hyperalgesia induced by CFA, as well as edema at the acute phase. Reduction of myeloperoxidase activity and inflammatory cell infiltration was observed in the paw tissue of mice injected with CFA and treated with EtOAc fraction. Together, our results support the antinociceptive and anti-inflammatory effects of the polyphenol-enriched fraction of A. crassiflora fruit peel and suggest that these effects are triggered, at least in part, by suppressing pro-inflammatory cytokines and neutrophils infiltration.

Effect of ionizing radiation after-therapy interval on bone: histomorphometric and biomechanical characteristics.

OBJECTIVES: This study aimed to evaluate the effects of radiotherapy on biomechanical, histomorphometric, and microstructural characteristics of bone, in diverse periods, compared with intact bone tissue. MATERIALS AND METHODS: Eighteen adult male New Zealand rabbits were treated with a single radiation dose of 30 Gy. The animals were randomly divided into six groups: NoIr, control group, no radiation, and five irradiated groups sacrificed after 24 h (Ir24h), 7 (Ir7d), 14 (Ir14d), 21 (Ir21d), and 28 (Ir28d) days. After these periods, the animals were sacrificed and their tibias (n = 6) evaluated using three-point bending test to calculate the ultimate force, work to failure, and bone stiffness. Dynamic indentation test was used to quantify Vickers hardness and elasticity modulus of bone tissue. Micro-CT was used to analyze the cortical volume (CtV), cortical thickness (CtTh), and porosity (Ct.Po). Histomorphometric assessment was based on the lacunarity of bone tissue. Data were analyzed using one-way ANOVA and Kruskal-Wallis tests followed by Tukey, Dunnet, and Dunn's post-tests (P < 0.05). RESULTS: The ultimate force, work to failure, stiffness, elastic modulus, and Vickers hardness values of irradiated bone were significantly lower that non-irradiated bone. Irradiated bone showed significantly lower CtTh and CtV values and higher CtPo than non-irradiated bone. No significant difference was found for lacunarity between non-irradiated bone and irradiated bone. CONCLUSIONS: Ionizing radiation decreases normal anisotropy on microarchitecture of cortical bone, and increases bone fragility compared with non-irradiated bone. Further, these changes were seen after longer periods (e.g., 14 and 21 days), and not immediately after radiation therapy. CLINICAL RELEVANCE: The radiotherapy reduces bone mechanical properties and the normal structure of organic and inorganic bone matrix. For studying the protocols to protect the radiotherapy effect using rabbit model, the use of the sacrificing period between 14 and 21 days is recommended.

Hyperbaric Oxygen therapy effects on bone regeneration in Type 1 diabetes mellitus in rats.

PURPOSE: The aim of this study was evaluate the effect of HBO on diabetic rats. MATERIALS AND METHODS: Twenty rats were distributed into four groups (n = 5): Control (C); Control + HBO (CH); Diabetes (D) and Diabetes + HBO (DH). Diabetes was induced by streptozotocin, and bone defects were created in both femurs in all animals. HBO therapy began immediately after surgery and was performed daily in the CH and DH groups. After 7 days, the animals were euthanized. The femurs were removed, demineralized, embedded in paraffin, and histologic images were analyzed. RESULTS: Qualitative histologic analyses showed more advanced stage bone regeneration in control groups (C and CH) compared with diabetic groups (D and DH). Histomorphometric analysis showed significantly increased bone neoformation in CH compared with the other groups (p < 0.001). Diabetic Group (D) showed decreased bone neoformation compared with non-diabetic groups (C and CH) (p < 0.001); however DH did not differ from C Group (p > 0.05). The mast cell population increased in CH compared with the other groups (C, D, and DH) (p < 0.05). The mast cell population did not differ between D and DH Groups. CONCLUSIONS: This study showed that HBO therapy improved early bone regeneration in diabetic rats and increased the mast cell population only in non-diabetic animals. HBO was shown to be important treatment for minimizing deleterious effects of diabetes on bone regeneration.

Influence of Platelet-Poor Plasma on Angiogenesis and Maintenance of Volume in Autogenous Bone Grafts.

PURPOSE: The aim of this study was to evaluate the effect of different storage media on angiogenesis and maintaining autogenous bone graft volume in rabbits. MATERIAL AND METHODS: Two grafts were removed bilaterally from the calvaria of 18 rabbits. One graft was removed and immediately fixed in the right mandibular angle (control group). The other graft was stored for 30 minutes in 1 of the following storage media (n = 6): saline solution (saline group), air exposure (dry group), or platelet-poor plasma (PPP group) and then retained by a screw in the right mandibular angle in the same animal. Four weeks later the animals were euthanized, and the grafted areas were harvested, fixed in 10% phosphate buffered formaldehyde solution, and embedded in paraffin. The 5-µm semi-serial sections were stained in hematoxylin and eosin and Mallory trichrome. RESULTS: Histologic analysis of all groups showed the bone graft was vascularized and well incorporated into the recipient site. The number of blood vessels decreased in the saline and dry groups compared with the control group (P < .03); in contrast, the number of blood vessels increased in the PPP group (P < .05). There were fewer osteoclasts in the saline group compared with the control group (P < .05). Furthermore, the saline group showed larger numbers of blood vessels than the dry group (P < .01). The PPP group showed larger bone graft volumes compared with the dry and saline groups (P < .01). In addition, the saline group showed larger bone graft volumes than the dry group (P < .01). CONCLUSIONS: PPP improved angiogenesis, maintained the volume of the autogenous bone graft, and was a better storage medium during the trans-surgical period than the dry and saline media.

The Effect of Local Delivery Doxycycline and Alendronate on Bone Repair.

The aim of the present study was to investigate the local effect of 10% doxycycline and 1% alendronate combined with poly(lactic-co-glycolic acid) (PLGA) on bone repair. Thirty rats were divided into three groups, as follows: control group (CG), drug group (DG), and vehicle-PLGA group (VG). Bone defect was created in the right femur and filled with the following: blood clot (CG); PLGA gel, 10% doxycycline and 1% alendronate (DG); or vehicle-PLGA (VG). The animals were euthanized 7 or 15 days after surgery. Bone density, bone matrix and number of osteoclasts were quantified. At 7 days, the findings showed increased density in DG (177.75 ± 76.5) compared with CG (80.37 ± 27.4), but no difference compared with VG (147.1 ± 41.5); no statistical difference in bone neoformation CG (25.6 ± 4.8), VG (27.8 ± 4), and DG (18.9 ± 7.8); and decrease osteoclasts in DG (4.6 ± 1.9) compared with CG (26.7 ± 7.4) and VG (17.3 ± 2.7). At 15 days, DG (405.1 ± 63.1) presented higher density than CG (213.2 ± 60.9) and VG (283.4 ± 85.8); there was a significant increase in percentage of bone neoformation in DG (31.5 ± 4.2) compared with CG (23 ± 4), but no difference compared with VG (25.1 ± 2.9). There was a decreased number of osteoclasts in DG (20.7 ± 4.7) and VG (29.5 ± 5.4) compared with CG (40 ± 9.4). The results suggest that the association of 10% doxycycline and 1% alendronate with PLGA-accelerated bone repair.

Low-level laser therapy on bone repair: is there any effect outside the irradiated field?

The biological effects of local therapy with laser on bone repair have been well demonstrated; however, this possible effect on bone repair outside the irradiated field has not been evaluated. The aim of this study was to investigate the effect of low-level laser therapy (LLLT) (λ = 830 nm) on repair of surgical bone defects outside the irradiated field, in rats. Sixty Wistar rats were submitted to osteotomy on the left femur and randomly separated into four groups (n = 15): group I, control, bone defect only; group II, laser applied on the right femur (distant dose); group III, laser applied locally on the bone defect and also on the right femur (local and distant doses); and group IV, laser applied locally on the left femur (local dose). Laser groups received applications within a 48-h interval in one point per session of density energy (DE) = 210 J/cm(2), P = 50 mW, t = 120 s, and beam diameter of 0.028 cm. Five animals of each group were euthanized 7, 15, and 21 days after surgery. Histologic analysis in all groups showed new bone formation in the region of interest (ROI) at 7 days. After 15 days, bone remodeling with a decrease of bone neoformation in the marrow area was observed in all groups. After 21 days, advanced bone remodeling with new bone mostly located in the cortical area was observed. The histomorphometric analysis showed at 7 days a significant increase of bone formation in groups III and IV compared to groups I and II. At days 15 and 21, histomorphometric analysis showed no significant differences between them. Laser therapy presented a positive local biostimulative effect in the early stage of bone healing, but the LLLT effect was not observed a long distance from the evaluated area.

Effect of low-level laser therapy on repair of the bone compromised by radiotherapy.

Radiotherapy (RDT) is commonly used for cancer treatment, but high doses of ionizing radiation can directly affect healthy tissues. Positive biological effects of low-level laser therapy (LLLT) on bone repair have been demonstrated; however, this effect on surgical defects of bone previously compromised by radiotherapy has not been evaluated. The aim of this study was to investigate the influence of LLLT (λ = 830 nm) in femur repair after ionizing radiation. Twenty Wistar rats were divided into four groups: control group (GC, n = 5) creation of bone defects (BDs) only; laser group (GL), with BD and LLLT (n = 5); radiotherapy group (GR), submitted to RDT and BD (n = 5); and radiotherapy and laser group (GRL), submitted to RDT, BD, and LLLT (n = 5). GL and GRL received punctual laser application (DE = 210 J/cm(2), P = 50 mW, t = 120 s, and beam diameter of 0.04 cm(2)) immediately after surgery, with 48-h interval during 7 days. Animals were euthanized at 7 days after surgery, and bone sections were evaluated morphometrically with conventional microscopy. Bone repair was only observed in nonirradiated bone, with significant improvement in GL in comparison to GC. GR and GRL did not present any bone neoformation. The result demonstrated a positive local biostimulative effect of LLLT in normal bone. However, LLLT was not able to revert the bone metabolic damage due to ionizing radiation.

Insulin Therapy on Bone Macroscopic, Microarchitecture, and MechanicalProperties of Tibia in Diabetic Rats.

BACKGROUND: This study evaluated tibia's macroscopic structure, mechanical properties, and bone microarchitecture in rats with type 1 diabetes mellitus (T1DM). METHODS: Eighteen animals were divided into three groups (n=6): Non-diabetic (ND), diabetic (D), and diabetic+insulin (DI). T1DM was induced by streptozotocin; insulin was administered daily (4IU). The animals were euthanized 35 days after induction. The tibiae were removed and analyzed using macroscopic, micro-computed tomography (micro-CT) and three-point bending. The macroscopic analysis measured proximal-distal length (PD), antero-posterior thickness (AP) of proximal (AP-P) and distal (AP-D) epiphysis, and lateral-medial thickness (LM) of proximal (LM-P) and distal (LM-D) epiphysis. Micro-CT analysis closed porosity, tissue mineral density, and cortical thickness. The three-point bending test measured maximum strength, energy, and stiffness. RESULTS: The macroscopic analysis showed that D presented smaller measures of length and thickness (AP and AP-P) than ND and DI. More extensive measurements were observed of LM and AP-D thickness in DI than in D. In micro-CT, DI showed larger cortical thickness than D. Mechanical analysis showed lower strength in D than in other groups. CONCLUSIONS: T1DM reduces bone growth and mechanical strength. Insulin therapy in diabetic rats improved bone growth and fracture resistance, making diabetic bone similar to normoglycemic animals.

Fibrin Network and Platelets Densities in Platelet-Rich Fibrin (PRF) Membranes Produced from Plastic Tubes Without Additives: A New Approach to PRF Clinical Use.

Background/Purpose: The present study aimed to investigate plastic tubes without additives as alternatives to glass and silica-coated plastic tubes, in the production of PRF membranes. Materials and Methods: Nine blood samples were collected from eight volunteers (n = 8) separated into three groups, according to tube material: glass, silica-coated plastic, and plastic without additives. In each group, the samples were centrifuged using different relative centrifugation forces: L-PRF (700 g/12 min), A-PRF (200 g/14 min), and A-PRF + (200 g/8 min). The generated membranes were evaluated by histomorphometry, considering the fibrin network, platelet aggregates, and cellular morphology, by light microscopy. The ultrastructural cellular morphology integrity was evaluated by transmission electron microscopy. Results: The L-PRF (p < 0.019) and A-PRF (p < 0.001) membranes showed a significantly lower fibrin network density in plastic tubes without additives compared to glass and silica-coated plastic tubes. Plastic tubes without additives revealed a significantly higher platelet percentage, regardless of the protocol (p < 0.005). In all groups, TEM analysis showed preserved normal morphological ultrastructure, maintaining the integrity of cellular components. Conclusion: Plastic tubes without additives offer a viable alternative for producing PRF membranes. They exhibited a higher platelet density and demonstrated fibrin network and cellular morphology similar to those of glass and silica-coated plastic tubes, irrespective of the centrifugation protocol.

Effect of different storage media on the regenerative potential of autogenous bone grafts: a histomorphometrical analysis in rabbits.

The success of autogenous bone graft is related to the graft cell viability. In bone-grafting procedures, harvested grafts are often maintained in extraoral media while the recipient site is prepared. The aim of this study was to evaluate in vivo the effect of storage media over autogenous bone grafts during the transsurgical time. Two grafts were removed bilaterally from the calvaria of 18 rabbits. One graft was immediately fixed in the mandibular angle (control group), and the other was maintained in air exposure (dry group), 0.9% NaCl solution (saline group), or platelet-poor plasma (PPP group) during 30 minutes and stabilized in the symmetrical location of control grafts. After 28 days, the animals were euthanized and the bone fragments were removed, demineralized, and embedded in paraffin. Histological evaluation was performed under light microscope. Empty lacunae and bone graft area quantification were carried out for the sections. The histomorphometrical analysis revealed reduction of the graft area and increase of empty lacunae in the dry group when compared with control. No significant differences were found in the number of empty lacunae or bone graft area between the saline group and its control and also between the PPP group and its control. The dry group showed more empty lacunae and less graft area than the saline and PPP groups. In accordance with the results, PPP and physiologic solution demonstrated osteocyte preservation and bone graft area maintenance, being satisfactory storage media for autogenous bone grafts during the transsurgical period.

Evaluation of the bone healing process utilizing platelet-rich plasma activated by thrombin and calcium chloride: a histologic study in rabbit calvaria.

To evaluate the bone healing of defects filled with particulate bone graft in combination with platelet-rich plasma (PRP), added with a mixture of calcium chloride and thrombin or just calcium chloride. Two 5-mm bone defects were created in the calvaria of 24 rabbits. Each defect was filled with particulate bone graft and PRP. In one defect the PRP was activated by a mixture of calcium chloride and thrombin; in the other, PRP was activated by calcium chloride only. The animals were euthanized 1, 2, 4, and 8 weeks after the surgeries, and the calvaria was submitted to histologic processing for histomorphometric analysis. The qualitative analysis has shown that both defects presented the same histologic characteristics so that a better organized, more mature, and well-vascularized bone tissue was noticed in the eighth week. A good bone repair was achieved using either the mixture of calcium chloride and thrombin or the calcium chloride alone as a restarting agent of the coagulation process.

Would nitric oxide be an effective marker for earlier stages of peri-implant disease? An analysis in human peri-implant sulcular fluid.

Nitric oxide has an important effect on host immune response. However, little has been studied in relation to its potential as a possible diagnostic tool in peri-implant disease. The present study analyzed nitrite levels in the peri-implant sulcular fluid (PISF) of implants with mucositis and the correlation of these nitrite levels with clinical parameters using a simplified fluid collection methodology. Twenty-five partially edentulous patients showing peri-implant mucositis were evaluated, and the peri-implant status was determined based on current clinical parameters: probing depth (PD) and bleeding on probing (BOP). The sulcular fluid (SF) around teeth (control) and implants were collected, and the nitrite levels were evaluated using the Griess method. The mean probing depth (mm) was significantly higher (P < .0001) in implants (2.852 ± 0.6484) than in control teeth (1.585 ± 0.3636). The mean total nitrite level (µM) was statistically higher (P = .0069) in implants with mucositis (14.34 ± 11.83) than in control teeth (9.316 ± 5.534). No correlation was observed between the total nitrite levels and the PD mean in the control group (P = .2558, r = -0.2361) or in the implant group (P = .1160, r = -0.3224), as well as the number of faces showing bleeding on probing (P = .8747, r = 0.0332). These results demonstrated that the nitrite levels were higher in inflamed areas. According to the methodology applied and results obtained, the higher nitrite levels in inflamed areas suggest that, in the future, nitrite could be used as a marker of peri-implant mucositis associated with clinical data to monitor the cure or evolution of the disease.

Progressive Platelet Rich Fibrin tissue regeneration matrix: Description of a novel, low cost and effective method for the treatment of chronic diabetic ulcers-Pilot study.

INTRODUCTION: Chronic lower limb ulcers (CLLU) are those injuries that persist for more than six weeks despite adequate care. They are relatively common; it is estimated that 10/1,000 people will develop CLLU in their lifetime. Diabetic ulcer, because of its unique pathophysiology (association between neuropathy, microangiopathy, and immune deficiency), is considered one of the most complex and difficult etiologies of CLLU for treatment. This treatment is complex, costly, and sometimes frustrating, as it is often ineffective, which worsens the quality of life of patients and makes its treatment a challenge. OBJECTIVE: To describe a new method for treating diabetic CLLU and the initial results of using a new autologous tissue regeneration matrix. METHOD: This is a pilot, prospective, an interventional study that used a novel protocol of autologous tissue regeneration matrix for the treatment of diabetic CLLU. RESULTS: Three male cases with a mean age of 54 years were included. A total of six Giant Pro PRF Membrane (GMPro) were used varying their application between one to three sessions during treatment. A total of 11 liquid phase infiltrations were performed varying their application between three and four sessions. The patients were evaluated weekly and a reduction in the wound area and scar retraction was observed during the period studied. CONCLUSION: The new tissue regeneration matrix described is an effective and low-cost method for the treatment of chronic diabetic ulcers.

Automated bone healing evaluation: New approach to histomorphometric analysis.

This study aimed to assess different approaches for bone healing evaluation on histological images and to introduce a new automatic evaluation method based on segmentation with distinct thresholds. We evaluated the hyperbaric oxygen therapy (HBO) effects on bone repair in type 1 diabetes mellitus rats. Twelve animals were divided into four groups (n = 3): non-diabetic, non-diabetic + HBO, diabetic, and diabetic + HBO. Diabetes was induced by intravenous administration of streptozotocin (50 mg/kg). Bone defects were created in femurs and HBO was immediately started at one session/day. After 7 days, the animals were euthanized, femurs were removed, demineralized, and embedded in paraffin. Histological sections were stained with hematoxylin and eosin (HE) and Mallory's trichrome (MT), and evaluated using three approaches: (1) conventional histomorphometric analysis (HE images) using a 144-point grid to quantify the bone matrix; (2) a semi-automatic method based on bone matrix segmentation to assess the bone matrix percentage (MT images); and (3) automatic approach, with the creation of a plug-in for ImageJ software. The time required to perform the analysis in each method was measured and subjected to Bland-Altman statistical analysis. All three methods were satisfactory for measuring bone formation and were not statistically different. The automatic approach reduced the working time compared to visual grid and semi-automated method (p < .01). Although histological evaluation of bone healing was performed successfully using all three methods, the novel automatic approach significantly shortened the time required for analysis and had high accuracy.

Type I Diabetes Mellitus and Insulin Therapy on Bone Microarchitecture, Composition and Mechanical Properties.

BACKGROUND: The aim of this study was to evaluate the microarchitecture, composition and mechanical properties of cortical bone of rats with type I diabetes mellitus (TIDM) and submitted to insulin therapy (IT). METHODS: Thirty rats were divided into three groups (n=10): non-diabetic, diabetic and diabetic+insulin. TIDM was induced by intravenous injection of streptozotocin. In diabetic+insulin group, 4IU insulin was administered twice per day (1I U at 7 am and 3I U at 7 pm). The animals were euthanized five weeks after TIDM induction; the tibiae were removed and submitted to microcomputed tomography (micro-CT, 8 µm), fourier transform infrared spectroscopy (FTIR) and dynamic microhardness indentation. RESULTS: Micro-CT analysis showed that diabetic group had lower bone surface/tissue volume ratio (BS/BV) (p=0.018), cortical thickness (Ct.Th) (p<0.001) and degree of anisotropy (Ct.DA) (p=0.034) values compared to non-diabetic group. The diabetic group showed lower Ct.Th than diabetic + insulin group (p=0.018). The non-diabetic group had lower fractal dimension (Ct.FD) values compared to diabetic groups (p<0.001). The ATR-FTIR analyses showed lower values for all measured parameters in the diabetic group than the non-diabetic group (amide I ratio: p=0.046; crystallinity index: p=0.038; matrix:mineral ratios - M:MI: p=0.006; M:MIII: p=0.028). The diabetic+ insulin group showed a lower crystallinity index (p=0.022) and M:MI ratio (p=0.002) than nondiabetic and diabetic groups, respectively. The diabetic group showed lower Vickers hardness values than non-diabetic (p<0.001) and diabetic+insulin (p=0.003) groups. CONCLUSION: TIDM negatively affects bone microarchitecture, collagen maturation, mineralization and bone microhardness. Moreover, insulin minimized the effect of TIDM on cortical thickness and organic/mineral matrix.