Recent refinements to cranial implants for rhesus macaques (Macaca mulatta).

The advent of cranial implants revolutionized primate neurophysiological research because they allow researchers to stably record neural activity from monkeys during active behavior. Cranial implants have improved over the years since their introduction, but chronic implants still increase the risk for medical complications including bacterial contamination and resultant infection, chronic inflammation, bone and tissue loss and complications related to the use of dental acrylic. These complications can lead to implant failure and early termination of study protocols. In an effort to reduce complications, we describe several refinements that have helped us improve cranial implants and the wellbeing of implanted primates.

Distribution and Morphology of Calcium-Binding Proteins Immunoreactive Neurons following Chronic Tungsten Multielectrode Implants.

The development of therapeutic approaches to improve the life quality of people suffering from different types of body paralysis is a current major medical challenge. Brain-machine interface (BMI) can potentially help reestablishing lost sensory and motor functions, allowing patients to use their own brain activity to restore sensorimotor control of paralyzed body parts. Chronic implants of multielectrodes, employed to record neural activity directly from the brain parenchyma, constitute the fundamental component of a BMI. However, before this technique may be effectively available to human clinical trials, it is essential to characterize its long-term impact on the nervous tissue in animal models. In the present study we evaluated how chronic implanted tungsten microelectrode arrays impact the distribution and morphology of interneurons reactive to calcium-binding proteins calbindin (CB), calretinin (CR) and parvalbumin (PV) across the rat's motor cortex. Our results revealed that chronic microelectrode arrays were well tolerated by the nervous tissue, with recordings remaining viable for up to 6 months after implantation. Furthermore, neither the morphology nor the distribution of inhibitory neurons were broadly impacted. Moreover, restricted microglial activation was observed on the implanted sites. On the whole, our results confirm and expand the notion that tungsten multielectrodes can be deemed as a feasible candidate to future human BMI studies.

Effect of propranolol on capsular reaction around silicone implants in guinea pigs.

PURPOSE: To evaluate the effect of propranolol on capsular architecture around silicone implants by measuring the inflammation, capsular thickness, and collagen fiber density, using a guinea pig experimental model. METHODS: Thirty six adult male guinea pigs randomly divided into two groups (n=18) were used. Each one received a silicone implant with textured-surface. The capsular tissue around implants from untreated or treated animals with the beta-adrenoceptor antagonist propranolol (10 mg/kg, dissolved in daily water) were analyzed for inflammation by histological scoring, capsular thickness by computerized histometry, and collagen fibers type I and Type III density by picrosirius polarization at different time points (7, 14 or 21 days after silicone implantation). RESULTS: Propranolol treatment reduced inflammation and impaired capsular thickness and delayed collagen maturation around the textured implant. CONCLUSION: Propranolol reduces the risk of developing capsular contracture around silicone implants with textured surface.

Effect of propranolol on capsular reaction around silicone implants in guinea pigs

PURPOSE: To evaluate the effect of propranolol on capsular architecture around silicone implants by measuring the inflammation, capsular thickness, and collagen fiber density, using a guinea pig experimental model. METHODS: Thirty six adult male guinea pigs randomly divided into two groups (n=18) were used. Each one received a silicone implant with textured-surface. The capsular tissue around implants from untreated or treated animals with the beta-adrenoceptor antagonist propranolol (10 mg/kg, dissolved in daily water) were analyzed for inflammation by histological scoring, capsular thickness by computerized histometry, and collagen fibers type I and Type III density by picrosirius polarization at different time points (7, 14 or 21 days after silicone implantation). RESULTS: Propranolol treatment reduced inflammation and impaired capsular thickness and delayed collagen maturation around the textured implant. CONCLUSION: Propranolol reduces the risk of developing capsular contracture around silicone implants with textured surface. .

Fractal dimension and Shannon's entropy analyses of the architectural complexity caused by the inflammatory reactions induced by highly crystalline poly(vinyl alcohol) microspheres implanted in subcutaneous tissues of the Wistar rats.

The results of the histopathological analyses after the implantation of highly crystalline PVA microspheres in subcutaneous tissues of Wistar rats are here in reported. Three different groups of PVA microparticles were systematically studied: highly crystalline, amorphous, and commercial ones. In addition to these experiments, complementary analyses of architectural complexity were performed using fractal dimension (FD), and Shannon's entropy (SE) concepts. The highly crystalline microspheres induced inflammatory reactions similar to the ones observed for the commercial ones, while the inflammatory reactions caused by the amorphous ones were less intense. Statistical analyses of the subcutaneous tissues of Wistar rats implanted with the highly crystalline microspheres resulted in FD and SE values significantly higher than the statistical parameters observed for the amorphous ones. The FD and SE parameters obtained for the subcutaneous tissues of Wistar rats implanted with crystalline and commercial microparticles were statistically similar. Briefly, the results indicated that the new highly crystalline microspheres had biocompatible behavior comparable to the commercial ones. In addition, statistical tools such as FD and SE analyses when combined with histopathological analyses can be useful tools to investigate the architectural complexity tissues caused by complex inflammatory reactions.

Alterations in the dynamics of inflammation, proliferation and apoptosis in subcutaneous implants of lupus-prone mice.

Wound repair is a complex process that involves inflammation, proliferation, extracellular matrix deposition/remodeling and apoptosis. Autoimmune diseases profoundly affect the healing process. We have used histological parameters to characterize the recruitment of mast cells and the proliferative activity and apoptosis in the fibrovascular tissue induced by subcutaneous polyether-polyurethane sponge implants in lupus-prone New Zealand White (NZW) and in control Balb/c mouse strains at days 10 and 21 post implantation. Fibrovascular tissue infiltration (hematoxylin and eosin staining), mast cell number (Dominici staining) and cellular proliferation (AgNOR staining) peaked early (day 10) but collagen deposition (picrosirius red staining) and apoptosis remained high in implants of NZW mice during the experimental period. In contrast, implants of Balb/c animals showed a progressive increase in mast cell recruitment and cellular proliferation but apoptosis fell from day 10 to 21 post-implantation. This divergent response early mast cells recruitment, excessive collagen deposition and disturbed removal of apoptotic cells from the site of injury in NZW mice implies that the genotype trait of NZW mice is a determining factor in abnormal healing response.

Resposta tecidual a implantes de discos de poliuretana de mamona nas formas pré-moldada ou biomassa moldada / Tissue reaction to premolded or molded biomass disks of polyurethanes containing castor oil

O trabalho teve por objetivo avaliar a resposta tecidual à implantação de discos de poliuretana derivada do óleo de mamona confeccionados de duas formas distintas, na forma pré-moldada fornecida pela indústria e em biomassa moldada no momento da aplicação, de forma a observar se haveria algum tipo de reação local tardia associada à possível continuidade do processo de endurecimento final do biomaterial. Foram utilizados 20 ratos, linhagem Wistar, fêmeas, com peso de 300-350g, nos quais foi inserido o disco pré-moldado no tecido subcutâneo do flanco direito e o de biomassa moldada no flanco esquerdo. Ambos os discos tinham 0,8cm de diâmetro por 0,5cm de espessura. Para o procedimento histológico, cinco ratos foram submetidos à eutanásia aos 3, 7, 15 e 30 dias de pós-cirúrgico. Os implantes e tecidos circundantes foram colhidos, processados e corados pela técnica de hematoxilina-eosina. Foi observada inicialmente uma reação inflamatória moderada, composta especialmente por células polimorfonucleares e macrófagos. Os linfócitos variaram de ausentes a discretos. A reação inflamatória diminui de intensidade à medida que se intensificou a formação de tecido conjuntivo fibroso ao redor dos implantes, porém sem modificação dos números de macrófagos. Sendo assim, conclui-se que ambos os discos de poliuretana induzem uma reação inflamatória similar, que varia de moderada a discreta na dependência do momento de avaliação.

The aim of this study was to histologically evaluate the tissue reaction to implantation of polyurethane disks containing castor oil produced in premolded form, supplied commercially, and biomass molded at the moment of application; since a late tissue reaction may occur during the total hardening process of the biomaterial. Twenty female Wistar rats, about 3 months of age, weighing 300-350g, and polyurethane disks 0.8cm in diameter and 0.5cm in thickness were used. The premolded disk was implanted into the subcutaneous tissue at the right flank and the molded biomass disk at the left flank. Five rats each were submitted to euthanasia at 3, 7, 15 and 30 days after surgery, and the polyurethane disks and surrounding tissue were collected. The specimens were processed for HE staining and examined microscopically. A moderate inflammatory reaction was primarily composed of polymorphonuclear cells and macrophages. The lymphocytes varied from absent to discreet. The intensity of the inflammatory reaction decreased at the same time the formation of fibrous conjunctive tissue increased around the implants. However, the numbers of macrophages remained the same. In conclusion, both polyurethane disks induce the same type of inflammatory reaction that varies from slight to moderate according to evaluation time.

Beta-adrenoceptor blockade delays granulation tissue formation in polyurethane sponge implants.

BACKGROUND: The role of adrenoceptors in granulation tissue formation is not well understood. The aim of this study was to investigate the effects of alpha- and beta-adrenoceptor blockade on granulation tissue development using polyurethane (PU) implants in the rat. METHODS: Animals were treated orally with propranolol (beta1- and beta2-antagonist), atenolol (beta1-antagonist) or phentolamine (alpha1- and alpha2-antagonist) until euthanasia. The control group received only water. All animals received subcutaneous implants of PU sponges. After 14 days, implants were collected, formalin-fixed and paraffin-embedded. Sections were stained with hematoxylin and eosin and Sirius red and immunostained for CD68 and alpha-smooth muscle actin. RESULTS: The number of inflammatory cells and the volume density of myofibroblasts and blood vessels were lower in the control group than in the propranolol- and atenolol-treated groups. The collagen fiber score was greater in the control group than in the propranolol- and atenolol-treated groups. The inflammatory infiltrate, collagen fiber score, blood vessel density or myofibroblast differentiation was not affected by phentolamine. The percentage of fibrovascular invasion was greater in the antagonist-treated groups than in the control group. CONCLUSIONS: Blockade of beta1- and beta2-adrenoceptors, but not alpha-adrenoceptors, impairs granulation tissue development in PU implants due to interference with the inflammatory response.

Pirfenidone prevents capsular contracture after mammary implantation.

BACKGROUND: Pirfenidone (PFD), a new antifibrotic and antiinflammatory agent, prevents and resolves fibrous tissue. This study evaluated the effect of PFD on adverse events in mammary implants using an animal model. Mammary implantation, the most frequent aesthetic surgery, may present several complications after surgery such as swelling, capsule contracture, hardness, and pain. METHODS: Wistar rats underwent submammary implantation with either smooth or textured silicone gel implants and were administrated 200 mg/kg of PFD daily. The control group received saline. The animals were killed at 8 weeks. The capsular tissue of both implants was removed for histologic and molecular analyses. RESULTS: Typical postaugmentation periimplant capsules with opacity on adjacent tissues developed 8 weeks after silicone implantation. No significant differences were observed between the textured and smooth implants in any analyzed parameter. Clearly, PFD reduced capsule thickness around submmamary tissue, fibroblast-like cell proliferation, and recruitment of inflammatory cells. The total cell numbers per field were reduced as well. In contrast, the control group presented abundant mononuclear cell infiltration and fibroblast-like cell proliferation. The total content of collagen in the PFD group was 50% less than in the control group. Fibroblast cells displayed 45% less activated phenotype in the PFD group than in the control group, as determined by immunohistochemistry techniques. In the PFD animals, transforming growth factor-beta (TGF-beta) decreased 85% and collagen 1 gene expression 60%, compared with the control group. CONCLUSION: The findings show a positive effect of PFD on mammary contracture in 10 rats. Despite the small number of animals, the differences found in 10 control rats encourage the authors to propose a larger study later and to suggest PFD as a potential preventive strategy in human mammary implantation surgery.

Implants of an adhesive resin, a calcium hydroxide, and a glass-ionomer cement cause reactional fibrosis with mast cells involvement in rats.

OBJECTIVES: To evaluate the tissue reaction of some pulpar protecting materials. METHOD AND MATERIALS: Standardized implants of calcium hydroxide (CH), glass-ionomer cement (GIC), and light-cured dentin adhesive (LDA), surgically introduced into the dorsal connective tissue of Wistar rats, were left for 15 and 30 days. For each of the 2 experimental times, a respective sham group (S) was studied. After experimentation, animals were sacrificed, and the material from implant sites was removed and studied with light microscopy and stereology (volume density of interstitial fibrosis, Vv[f], and numerical density of mast cells per area, NA[mast cell]). The implants and the surfaces of the fibrous capsule were analyzed with scanning electronic microscopy (SEM) adapted with microanalysis. RESULTS: The CH group had the smallest value of Vv[f] and the LDA group the greatest. At day 30, the Vv[f] of the LDA group showed an increase of 30% and was different from that of the CH group in the same period (P <.05). The NA[mast cell] was smaller in both the CH and the S group than in the LDA group (P <.05). The GIC group had intermediate values for Vv[f] and NA[mast cell] between CH and LDA values. There was light fibrosis in the surgical area with few mast cells associated to vessels in the S group. SEM detected presence of silica fragments in the fibrous capsule of the LDA group and calcium in the fibrous capsule of the CH group. CONCLUSIONS: All tested materials allowed healing of the implanted area; tissue reaction was smallest in the CH group.

Cellular proliferation, differentiation and apoptosis in polyether-polyurethane sponge implant model in mice.

The integration of implanted material to host organism requires spatial and temporal organization of several cellular processes, such as proliferation, differentiation and apoptosis. Despite the clinical relevance of these processes, there is little information regarding the sequence of such events in synthetic matrices. Here, we present a combination of techniques used to characterize the fibrovascular response in subcutaneous polyether-polyurethane sponge implants in mice at days 4, 7, 10 and 14 postimplantation. The AgNOR technique was modified and used as a surrogate marker for proliferating and activated cells invading the implant. The number of AgNOR-stained cells increased progressively from day 4 (606+/-76) to day 14 (2146+/-71) postimplantation. The number of TUNEL-positive (apoptotic index) cells also increased progressively from day 4 (459+/-40) to day 14 (1157+/-119) postimplantation. However, the ratio of TUNEL-labeled/proliferating cells had its highest peak in the early phase of the process remaining stable until day 14. Using Picrosirius staining it was shown that thin collagen increased from day 4, peaking at day 10 and falling markedly at day 14, whereas dense collagen increased progressively during the whole period. These experiments hold potential to investigate not only distinct phases of tissue repair induced by synthetic matrices but also to study underlying mechanisms involved.

Experimental peri-implant tissue breakdown around different dental implant surfaces: clinical and radiographic evaluation in dogs.

PURPOSE: Tissue reactions to 4 different implant surfaces were evaluated in regard to the development and progression of ligature-induced peri-implantitis. MATERIALS AND METHODS: In 6 male mongrel dogs, a total of 36 dental implants with different surfaces (9 titanium plasma-sprayed, 9 hydroxyapatite-coated, 9 acid-etched, and 9 commercially pure titanium) were placed 3 months after mandibular premolar extraction. After 3 months with optimal plaque control, abutment connection was performed. Forty-five days later, cotton ligatures were placed around the implants to induce peri-implantitis. At baseline and 20, 40, and 60 days after placement, the presence of plaque, peri-implant mucosal redness, bleeding on probing, probing depth, clinical attachment loss, mobility, vertical bone loss, and horizontal bone loss were assessed. RESULTS: The results did not show significant differences among the surfaces for any parameter during the study (P > .05). All surfaces were equally susceptible to ligature-induced peri-implantitis over time (P < .001). Correlation analysis revealed a statistically significant relationship between width of keratinized tissue and vertical bone loss (r2 = 0.81; P = .014) and between mobility and vertical bone loss (r2 = 0.66; P = .04), both for the titanium plasma-sprayed surface. DISCUSSION AND CONCLUSIONS: The present data suggest that all surfaces were equally susceptible to experimental peri-implantitis after a 60-day period.

Mechanisms of impaired vascular response to ANG II in perivascular injured carotid arteries of ovariectomized rat.

The rabbit carotid artery, injured by silicone collar, presents a perivascular inflammatory response and alterations in vascular responsiveness. Considering that angiotensin II (Ang II) plays an important role in cardiovascular physiology and pathology and that cardiovascular disease increases in postmenopausal women, the aim of this study was to investigate whether the Ang II contractile response in ovariectomized rat carotid artery is modified after a vascular injury by silicone collar. The positioning of the silicone collar around the common carotid artery for 14 days leads to an increased cross-sectional area of adventitial layer with inflammatory cells and an extensive angiogenesis. The Ang II-induced contraction was significantly decreased in collared arteries when compared with contralateral arteries. The reduction in the constrictor effect of Ang II in collared arteries was not modified by the presence of indomethacin (a non-selective inhibitor of cyclooxygenase) or PD 123,319 (a selective antagonist of the Ang II AT2 receptor). Moreover, while endothelium removal induced an increase in the Ang II responsiveness of both arteries (collared and contralateral), the Emax induced by Ang II was still lower in collared arteries. However, the "in vitro" pretreatment of the arteries with an inhibitor of nitric oxide synthase enzyme (L-NAME) significantly enhanced the maximal contractions response to Ang II only in injured arteries. Furthermore, the expression of iNOS (inducible nitric oxide synthase) was observed in the adventitial layer of collared arteries, indicating that the NO formed in the adventitial layer has an important role in injured arteries. Moreover, our data show impairment of extracellular calcium mobilization, mediated by Ang II, in the collared artery, although the intracellular calcium mobilization was not modified by the injury. In conclusion, the increased production of NO and a decrease in the calcium influx displayed by Ang II in the collared artery appears to counteract and reduce the biologic effect of Ang II.

Sponge-induced angiogenesis and inflammation in PAF receptor-deficient mice (PAFR-KO).

1. To determine biological functions of platelet-activating factor (PAF) in chronic inflammation, we have investigated the kinetics of angiogenesis, inflammatory cells recruitment and cytokine production in sponge-induced granuloma in wild type and PAF receptor-deficient mice (PAFR-KO). 2. Angiogenesis as determined by morphometric analysis and hemoglobin content was significantly higher in the implants of PAFR-KO mice at all time points. Treatment with PAF receptor antagonist UK74505 (30 mg kg(-1)) also increased angiogenesis in sponge implants. 3. Neutrophils and macrophages accumulation, as determined by myeloperoxidase and N-acetylglucosaminidase activities in the supernatant of implanted sponges were markedly decreased in PAFR-KO mice. Surprisingly, the levels of the proinflammatory chemokines, keratinocyte-derived chemokine and chemokine monocyte chemoattractant protein 1 were higher in the implants of the transgenic animals. 4. We have shown that angiogenesis was stimulated in PAFR-KO mice whereas inflammation was decreased, indicating that PAF is an endogenous regulator of new blood vessels formation in the inflammatory microenvironment induced by the sponge implant.

Clinical study of guided bone regeneration and/or bone grafts in the treatment of ligature-induced peri-implantitis defects in dogs.

This study evaluated, by clinical analysis, the hard tissue response following treatment of ligature-induced peri-implantitis defects in 5 dogs. The mandibular premolars were removed from both sides of the jaw. After 3 months of healing, two titanium implants were placed on each side of the mandible. Following abutment connection, 3 months later, experimental peri-implantitis was induced by the placement of cotton ligatures in a submarginal position. Ligatures and abutments were removed after one month and the bony defects were randomly assigned to one of the following treatments: debridement (DE), debridement plus guided bone regeneration (GBR), debridement plus mineralized bone graft (BG) and debridement plus guided bone regeneration associated with mineralized bone graft (GBR + BG). The peri-implant bone defects were clinically measured before and 5 months post-treatment. Results showed a higher percentage of vertical bone fill for GBR + BG (27.77 +/- 14.07) followed by GBR (21.78 +/- 16.19), BG (21.26 +/- 6.87), DE (14.03 +/- 5.6). However, there were no statistically significant differences between any of the treatments proposed (one way repeated measures analysis of variance, P = 0.265).

Absorbable versus nonabsorbable membranes and bone grafts in the treatment of ligature-induced peri-implantitis defects in dogs. Part I. A clinical investigation.

The purpose of this study was to clinically evaluate an absorbable collagen membrane (Bio-Gide) and a nonabsorbable polytetrafluoroethylene membrane (PTFE), associated or not with bone grafts, for the treatment of ligature-induced peri-implantitis defects in dogs. The bilateral mandibular premolars were removed from 5 2-year-old mongrel dogs. After 3 months of healing, 3 titanium implants were placed on each side of the mandible. Experimental peri-implantitis was induced after abutment connection. Ligatures and abutments were removed after 1 month and the bone defects were randomly assigned to one of the following treatments: DB: debridement alone; GBR+BG-I: debridement plus PTFE membrane associated with mineralized bone graft (Bio-Oss); GBR+BG-II: debridement plus collagen membrane (Bio-Gide) associated with mineralized bone graft; GBR-I: debridement plus PTFE membrane; GBR-II: debridement plus collagen membrane; BG: debridement plus mineralized bone graft. The peri-implant bone defects were measured before and 5 months after treatment. Results showed the greatest percentage of vertical bone fill for GBR+BG-II (27.77+/-14.07) followed by GBR-II (21.78+/-16.19), BG (21.26+/-6.87), GBR+BG-I (19.57+/-13.36), GBR-I (18.86+/-10.63) and DB (14.03+/-5.6). However, the values were not statistically significant (ANOVA, contrast F test, P=0.612). Within the limits of the present investigation, it can be concluded that no difference was detected among treatments.

Evaluation of guided bone regeneration and/or bone grafts in the treatment of ligature-induced peri-implantitis defects: a morphometric study in dogs.

The goal of this study was to evaluate, morphometrically, hard-tissue healing following the treatment of ligature-induced peri-implantitis defects in dogs and guided bone regeneration and/or bone grafts. Five dogs were used, and the mandibular premolars were removed. Three months later, two titanium implants were installed on each side of the mandible, and after another 3 months, abutment connection was performed. Following abutment connection, experimental periimplantitis was induced by placing cotton ligatures in a submarginal position. Ligatures and abutments were removed after 1 month and the bony defects were randomly assigned to one of the following treatments: debridement (DE), debridement plus guided bone regeneration (GBR), debridement plus mineralized bone graft (BG), and debridement plus guided bone regeneration associated with mineralized bone graft (GBR/BG). The dogs were euthanatized after 5 months. Morphometric analysis did not reveal significant differences among the treatments neither with respect to the percentage of bone to implant contact (p = 0.996) nor to the bone area (p = 0.946) within the limits of the threads of the implant. Within the limits of this investigation, there is insufficient evidence to indicate that any of the treatments presented an improved response in dealing with bony defects resulting from peri-implantitis.