PARTICIPATION OF MACROPHAGES IN CHLORAMPHENICOL-POTENTIATED CARRAGEENAN-INDUCED PERITONITIS IN RATS

1. We investigated the possible potentiating effect of chloramphenicol succinate (30 mg/kg, every 12 h for 4 days, ip) on the response of polymorphonuclear neutrophils to carrageenin (150 mug, ip) or dextran (100 mug, ip) in the peritoneal cavity of male Wistar rats (180-230 g; N = 12 in each group).2. Chloramphenicol potentiated the cell migration induced by carrageenin (35%) but not that induced by dextran. Previous macrophage depletion in the peritoneal cavity by washing with sterile saline abolished the cell response, whereas a previous thioglycollate-induced increase in macrophage numbers enhanced the potentiating effect (60%).3. These results suggest that the potentiating effect on polymorphonuclear neutrophil migration induced by chloramphenicol may be related to chemotactic factors released by macrophages.

Modulation of intercellular communication in macrophages: possible interactions between GAP junctions and P2 receptors

Gap junctions are connexin-formed channels that play an important role in intercellular communication in most cell types. In the immune system, specifically in macrophages, the expression of connexins and the establishment of functional gap junctions are still controversial issues. Macrophages express P2X(7) receptors that, once activated by the binding of extracellular ATP, lead to the opening of transmembrane pores permeable to molecules of up to 900 Da. There is evidence suggesting an interplay between gap junctions and P2 receptors in different cell systems. Thus, we used ATP-sensitive and -insensitive J774.G8 macrophage cell lines to investigate this interplay. To study junctional communication in J774-macrophage-like cells, we assessed cell-to-cell communication by microinjecting Lucifer Yellow. Confluent cultures of ATP-sensitive J774 cells (ATP-s cells) are coupled, whereas ATP-insensitive J774 cells (ATP-i cells), derived by overexposing J774 cells to extracellular ATP until they do not display the phenomenon of ATP-induced permeabilization, are essentially uncoupled. Western-blot and reverse-transcription polymerase chain reaction assays revealed that ATP-s and ATP-i cells express connexin43 (Cx43), whereas only ATP-s cells express the P2X(7) receptor. Accordingly, ATP-i cells did not display any detectable ATP-induced current under whole-cell patch-clamp recordings. Using immunofluorescence microscopy, Cx43 reactivity was found at the cell surface and in regions of cell-cell contact of ATP-s cells, whereas, in ATP-i cells, Cx43 immunoreactivity was only present in cytosolic compartments. Using confocal microscopy, it is shown here that, in ATP-s cells as well as in peritoneal macrophages, Cx43 and P2X(7) receptors are co-localized to the membrane of ATP-s cells and peritoneal macrophages.

Does alveolar soft-part sarcoma exhibit skeletal muscle differentiation? An immunocytochemical and biochemical study of myogenic regulatory protein expression

There has been persistent controversy regarding the nature of cell differentiation in alveolar soft-part sarcoma (ASPS) since its first description in 1952. Some studies suggest that ASPS might represent an unusual variant of skeletal muscle tumor, Given the availability of new monoclonal antibodies to probe for skeletal muscle differentiation and the rapid advance in immunocytochemical techniques for deparaffinized, formalin-fixed tissue sections, we wished to test the proposed hypothesis that ASPS might represent a new type of rhabdomyosarcoma Twelve archival samples of ASPS were retrieved, and we investigated the expression of two myogenic regulatory proteins, MyoD1 and myogenin, as rvell as other muscle-associated proteins, using sensitive immunocytochemical techniques. Despite the presence of desmin immunostaining in six ASPSs, no tumors were positive for either muscle actin or myoglobin Most importantly, no specimen showed nuclear expression of MyoD1 or myogenin, In 11 tumors, however, there was considerable granular immunostaining in the tumor cell cytoplasm with the anti-MyoD1 monoclonal antibody 5.8A, a phenomenon observed in various nonmuscle normal and neoplastic tissues with this antibody, To analyze the exact nature of immunostaining of MyoD1 and desmin in ASPS, biochemical analyses using available fresh frozen tumor tissue were performed, Although a 53-kDa band was noted with antidesmin antibody on Western blot analysis, no specific protein band that corresponds to the 45-kDa MyoD1 was detected with antibody 5.8A. These results confirm the presence of desmin in ASPS but argue against authentic expression of MyoD1, They also suggest that the cytoplasmic immunostaining observed with anti-MyoD1 antibody 5.8A most likely represents a nonspecific cross-reaction with an unknown cytoplasmic antigen, Considering the master role that MyoD1 and myogenin play in skeletal muscle commitment and differentiation and the lack of expression of these two proteins in ASPS as determined immunocytochemically and biochemically, we think that the histogenesis of ASPS remains unknown.

Osteoblasts engulf apoptotic bodies during alveolar bone formation in the rat maxilla

During bone formation, as in other tissues and organs, intense cellular proliferation and differentiation are usually observed. It has been described that programmed cell death, i.e., apoptosis, takes place in the control of the cellular population by removing of the excessive and damaged cells. Although it is generally accepted that apoptotic bodies are engulfed by professional phagocytes, the neighboring cells can also take part in the removal of apoptotic bodies. In the present study, regions of initial alveolar bone formation of rat molars were examined with the aim to verify whether osteoblasts are capable of engulfing apoptotic bodies, such as professional phagocytes. Rats aged 11-19 days were sacrificed and the maxillary fragments containing the first molar were removed and immersed in the fixative solution. The specimens fixed in glutaraldehyde-formaldehyde were processed for light microscopy and transmission electron microscopy. For the detection of apoptosis, the specimens were fixed in formaldehyde, embedded in paraffin, and submitted to the TUNEL method. The results revealed round/ovoid structures containing dense bodies on the bone surface in close contact to osteoblasts and in conspicuous osteoblast vacuoles. These round/ovoid structures showed also positivity to the TUNEL method, indicating that bone cells on the bone surface are undergoing apoptosis. Ultrathin sections showed images of apoptotic bodies being engulfed by osteoblasts. Occasionally, the osteoblasts exhibited large vacuoles containing blocks of condensed chromatin and remnants of organelles. Thus, these images suggest that osteoblasts are able to engulf and degrade apoptotic bodies. (c) 2005 Wiley-Liss, Inc.

Inferior alveolar nerve transposition with involvement of the mental foramen for implant placement

Inferior Alveolar Nerve (IAN) transposition is an option for prosthetic rehabilitation in cases of moderate or even severe bone reabsorption for patients that do not tolerate removable dentures. The aim of the present report is to describe an inferior alveolar nerve transposition with involvement of the mental foramen for implant placement. The surgical procedure was performed under local anesthesia, by the inferior alveolar, lingual and buccal nerve blocking technique. Centripetal osteotomy was performed, and bone tissue was removed, leaving the nerve tissue free in the foramen area. After that, transsection of the incisor nerve was performed, and lateral osteotomy was started from the buccal direction, toward the trajectory of the IAN. The procedure was concluded, by making use of a delicate resin spatula to manipulate the vascular-nervous bundle. The drilling sequence for placing the dental implants was performed, and autogenous bone was harvested using a bone collector attached to the surgical suction appliance. After the implants were placed, the bone tissue previously collected during the osteotomies and drilling processes was placed in order to protect the IAN from contact with the implants. The surgical protocol for inferior alveolar nerve transposition, followed by implant placement presented excellent results, with complete recovery of the sensitivity, seven months after the surgical procedure.

Alveolar process preservation at implants installed immediately into extraction sockets using deproteinized bovine bone mineral - an experimental study in dogs

Aim To evaluate the soft tissue and the dimensional changes of the alveolar bony crest at sites where deproteinized bovine bone mineral (DBBM) particles, concomitantly with the placement of a collagen membrane, were used at implants installed into sockets immediately after tooth extraction. Material and methods The pulp tissue of the mesial roots of 3P3 was removed in six Labrador dogs, and the root canals were filled. Flaps were elevated bilaterally, the premolars hemi-sectioned, and the distal roots removed. Recipient sites were prepared in the distal alveolus, and implants were placed. At the test sites, DBBM particles were placed in the residual marginal defects concomitantly with the placement of a collagen membrane. No treatment augmentation was performed at the control sites. A non-submerged healing was allowed. Impressions were obtained at baseline and at the time of sacrifice performed 4 months after surgery. The cast models obtained were analyzed using an optical system to evaluate dimensional variations. Block sections of the implant sites were obtained for histological processing and soft tissue assessments. Results After 4 months of healing, no differences in soft tissue dimensions were found between the test and control sites based on the histological assessments. The location of the soft tissue at the buccal aspect was, however, more coronal at the test compared with the control sites (1.8 +/- 0.8 and 0.9 +/- 0.8 mm, respectively). At the three-dimensional evaluation, the margin of the soft tissues at the buccal aspect appeared to be located more apically and lingually. The vertical dislocation was 1 +/- 0.6 and 2.7 +/- 0.5 mm at the test and control sites, respectively. The area of the buccal shrinkage of the alveolar crest was significantly smaller at the test sites (5.9 +/- 2.4 mm2) compared with the control sites (11.5 +/- 1.7 mm2). Conclusion The use of DBBM particles concomitantly with the application of a collagen membrane used at implants placed into sockets immediately after tooth extraction contributed to the preservation of the alveolar process.

Effects of remifentanil on the minimum alveolar concentration of isoflurane in dogs

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

COX-2 inhibition decreases VEGF expression and alveolar bone loss during the progression of experimental periodontitis in rats

Background: Vascular endothelial growth factor (VEGF) is a macromolecule of importance in inflammation that has been implicated in periodontitis. The aims of this study were to investigate VEGF expression during the progression of periodontal disease and to evaluate the effect of a preferential cyclooxygenase (COX)-2 inhibitor meloxicam on VEGF expression and alveolar bone loss in experimentally induced periodontitis.Methods: A total of 120 Wistar rats were randomly separated into groups 1 (control) and 2 (meloxicam, 3 mg/kg/day, intraperitoneally, for 3, 7, 14, or 30 days). Silk ligatures were placed at the gingival margin level of the lower right first molar of all rats. VEGF expression was assessed by reverse transcription-polymerase chain reaction (RT-PCR), Western blot (WB), and immunohistochemical (IHC) analyses. The hemiarcades were processed for histopathologic analysis. RT-PCR and WB results were submitted to analysis of variance, the Tukey test, and Pearson correlation analysis (P<0.05).Results: A reduction in alveolar bone resorption was observed in the meloxicam-treated group compared to the control group at all periods studied. There was a positive correlation between COX-2 mRNA and VEGF mRNA in the gingival tissues and periodontal disease (R = 0.80; P = 0.026). Meloxicam significantly reduced the increased mRNA VEGF expression in diseased tissues after 14 days of treatment (P = 0.023). Some alterations in VEGF receptor I mRNA expression were observed, but these were not statistically significant. VEGF protein expression in WB experiments was significantly higher in diseased sites compared to healthy sites (P<0.05). After 14 days of treatment with meloxicam, an important decrease in VEGF protein expression was detected in diseased tissues (P = 0.08). Qualitative IHC analysis revealed that VEGF protein expression was higher in diseased tissues and decreased in tissues from rats treated with meloxicam.Conclusions: The present data suggest an important role for VEGF in the progression of periodontal disease. Systemic therapy with meloxicam can modify the progression of experimentally induced periodontitis in rats by reducing VEGF expression and alveolar bone loss.

Comparison of the effects of epidural or intravenous methadone on the minimum alveolar concentration of isoflurane in dogs

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Renal interstitial foam cells are macrophages

Immunohistochemical studies on renal biopsies from eight patients with various types of glomerulonephritis showed that the interstitial foam cells belonged to the monocyte-macrophage lineage. There was a strong association between hypercholesterolaemia and the presence of renal interstitial foam cells.

Effects of fibrin adhesive material (Tissucol) on alveolar healing in rats under stress.

The effects of Tissucol on alveolar healing following stress were evaluated histologically, comparing three groups of 28 male albino rats each. Stress was applied and their right upper incisors were extracted. Group A served as an empty control site. In Group B, Tissucol was applied into the alveolar cavity. Group C received local antifibrinolytic treatment (alveolar irrigation with epsilon-aminocaproic acid solution) before implant of Tissucol into the tooth socket. Four animals in each group were killed at 1, 3, 6, 9, 15, 21 and 24 days after surgery. Results showed that: 1) Tissucol did not interfere with connective and osseous tissue formation; 2) Tissucol allowed new bone formation; 3) Tissucol residues in Group B in sections of 24-day specimens did not impair healing; 4) Tissucol was usually completely resorbed and healing was complete 24 days after surgery in Group C.

REDUCAO DA CONCENTRACAO ALVEOLAR MINIMA DO ISOFLURANO E DO SEVOFLURANO PELO SUFENTANIL

Background and Objectives - Inhalational anesthetics have a mild analgesic effect. The reduction of alveolar concentration (MAC) of potent volatile anesthesics by increasing plasma concentrations of opioids is desired in inhalational anesthesia. The purpose of this study was to determine the role of sufentanil in reducing sevoflurane and isoflurane MAC. Methods - Thirty eight adult patients of both genders, physical status ASA I or II, submitted to major abdominal procedures were randomly allocated into two groups. Group I (n = 24) received inahalational anesthesia with sevoflurane and Group II (n = 14) received inhalational anesthesia with isoflurane, both diluted in a mixture of N2O (1 liter) and O2 (0.5 liter). A semi-closed system with CO2 absorber and partial reinhalation was used. Ventilation was mechanically controlled. Sufentanil infusion was administered aiming at obtaining 0.5 ng.ml-1 of plasma concentration. Sufentanil plasma concentration was previously calculated by a computer software. End-tidal concentrations were obtained through a gas analyzer and measured at 15 minutes (M1), 30 minutes (M2), 60 minutes (M3), 90 minutes (M4) and 120 minutes (M5). Systolic and diastolic blood pressure (SBP and DBP) and heart rate (RR) were measured during the same periods with the addition of M0 (pre-anesthetic period). Hourly consumption of the inhalational anesthetic agent (IAC), extubation time (ET = time between admission to the recovery room and extubation) and stay in the post anesthesia recovery room (PA-RR) were also measured. Results - Type and duration of surgeries were similar for both groups. There were no statistically significant differences in MAC, SBP, DBP, RR, IAC, TE and PA-RR between groups. Systolic blood pressure in group I (sevoflurane) showed differences among periods F = 3.82 p < O.05; (M2 = M3)(M4 = M5) and M1 had a intermediate value. MAC in group I showed differences among periods F = 9.0 p < 0.05; M1 < M3. MAC in group II also showed differences among periods F = 13.03 p < O.05; M1 < (M2,M3,M4,M5). Conclusions - Both groups had similar behavior when associated to sufentanil in major abdominal surgeries. Group II showed a higher cardiac and circulatory stability.

Involvement of the major glycoprotein (gp43) of Paracoccidioides brasiliensis in attachment to macrophages

The yeast form of Paracoccidioides brasiliensis, the causative agent of a deep mycosis in humans, is known to be phagocytized by, and to multiply inside, macrophages. In this work we describe the involvement of gp43, a major antigenic protein of P. brasiliensis, in the initial steps of attachment of the fungus to macrophages. Anti-gp43 F(ab) polyclonal fragments were capable of inhibiting phagocytosis in a concentrationdependent manner. Sheep red blood cells sensitized with purified gp43 were more endocytized than SRBC alone, and this process was also inhibited by anti-gp43 F(ab) fragments. Inhibition tests indicated the involvement of fucose and mannose residues in the phagocytosis of the fungus and of SRBC-gp43 by macrophages. Taken together, these results suggest that gp43 may be involved in the adherence and uptake of the fungus by murine peritoneal macrophages, and that this binding may be dependent on monosaccharide residues that are part of the gp43 glycoprotein. © 1998 ISHAM.

Foamy macrophages in the liver of cattle fed Brachiaria brizantha hay

Liver and lymph nodes injuries characterized by clusters of foamy macrophages, some of them containing birefringent crystals, were observed in cattle fed on Brachiaria brizantha hay. The cattle were from an experimental group poisoned with Senecio brasiliensis known to cause hepatic fibrosis and hepatocyte megalocytosis. One of the animals developed photosensitivity but the exact cause wasn't determined since both plants were fed. The foamy macrophages were present from the 30th d of feeding. Early appearance of these lesions may be particular to the animal specie used or due to the presence of both toxic plants.

Influence of age on combined effects of cyclosporin and nifedipine on rat alveolar bone

Background: There is some evidence showing that cyclosporin A (CsA) and nifedipine (NIF) affect bone metabolism. The purpose of this work was to study the effects of CsA and NIF, given alone or concurrently, on alveolar bone of rats of different ages. Methods: Rats 15, 30, 60, and 90 days old were treated daily with 10 mg/kg body weight of CsA subcutaneously injected and/or 50 mg/kg body weight of NIF/day given orally for 60 days. Alveolar bone of the first lower molars was morphologically and stereologically evaluated in serial 5 μm bucco-lingual paraffin sections, stained with hematoxylin and eosin. Serum calcium and alkaline phosphatase levels were measured in all animals at the end of the experimental period. Results: Rats treated with CsA or NIF alone or CsA and NIF concurrently showed decreased alveolar bone density. CsA was more effective than NIF. A significant decrease in serum calcium was found only in animals treated with CsA or CsA/NIF. The results were similar regardless of age. Conclusions: These results indicate that the decrease in the alveolar bone volume in rats caused by CsA and NIF alone or concurrently is not age dependent. Furthermore, NIF (50 mg/kg) did not further increase the loss of alveolar bone volume induced by CsA (10 mg/kg).