Marlene Isabel Vargas Viloria

Association of mesenchymal stem cells with platelet rich plasma on the repair of critical calvarial defects in mice

PURPOSE To evaluate the effects of mesenchymal stem cells (MSC) from eight mice C57BL/6 gfp(+) bone marrows expanded in cultures associated with platelets rich plasma (PRP) deriving from another eight mice, in the repair of critical defects in calvarial bone produced in twenty-four adult isogenic mice C57BL/6. METHODS The animals were submitted to a cranial defect of 6.0mm in diameter and divided into two equal experimental groups. Control group did not receive treatment and the treated group received a MSC pellet containing 1.0 x 10(7) cells/mL associated with 50.0 µL of plasma gel containing 1.0 x 10(9) autologous platelets within the defect. RESULTS In the treated group was observed process of angiogenesis and bone repair better than control group. CONCLUSION Mesenchymal stem cells derived from bone marrow of C57BL/6 gfp(+) mice associated with PRP gel applied in bone critical defects produced in calvarial contributes positively to the process of bone repair.

Plasma rico em plaquetas associado ou não ao osso esponjoso autógeno no reparo de falhas ósseas experimentais

The present study evaluated autogenous platelet rich plasmas (PRP) influence on the reparation process of four bone defects made on rabbits skull, associated or not to autogenous bone graft (EOE). Defect I received PRP only; defect II received 3mg of EOE only; defect III received EOE associated to PRP; defect IV was left to heal naturally, serving as control. After each surgery the animals were randomly divided into three groups that were euthanized at 30, 60 and 90 days. In the mesoscopic evaluation bone ingrowth started from the defects borders to the center and from the bottom to the surface for all observation times on the control (VI) and PRP only (I) groups. In the groups treated with EOE only (II) and EOE associated to PRP (III) new bone was observed in the center of the defects. Radiographic analysis showed greater central radiopacity for groups treated with EOE only (II) and EOE associated to PRP (III) at all observation times. Microscopically in the group treated with EOE associated to PRP (III) at 30 days the graft was indistinguishable from new bone present on the border of the defect, associated to a moderate quantity of a very vascularized and cellular fibrous connective tissue. This tissue showed an extracelular eosinophilic amorphous foamy material, associated to an inflammatory process constituted by lymphocytes and in less number by macrophages and multinucleated giant cells that may have negatively influenced early bone formation. At 60 and 90 days occasional spots of lymphocytic inflammation were observed. Both treatments, PRP associated or not to EOE, were similar for the bone ingrowth at the final time of observation; the graft used alone determined early bone reparation and thromboplastine used for the platelet gel formation incited a foreign body-like reaction that acted negatively on the initial reparation.

Hidroxiapatita sintética pura, hidroxiapatita sintética associada ao colágeno e hidroxiapatita sintética associada ao lipossoma como substitutos ósseos em defeitos provocados na tíbia de cães: aspectos da osteointegração à microscopia de luz transmitida

The objective of this work was to evaluate histologically the synthetic hydroxyapatite (HAP-91), associated to collagen (COL.HAP-91) and associated to liposome (INT.HAP-91), as bone substitutes. Fourty clinically healthy dogs were studied. After a conventional anaesthesic protocol, the surgical procedure consisted of skin incision on medial surface in the left tibial third middle. A bone fragment of 10 ´ 6mm in size was retreated in all animals, which were divided into four groups with 10 animals each. Group one was treated with HAP-91, group two with COL.HAP-91, group three with INT.HAP-91, group four did not receive any treatment and was used as a control. Histological material was collected after humane sacrifice of two animals from each group in 8, 30, 60, 120 180 days after surgery. By the 8th day, bone neoformation in control group and around the implant in groups treated with HAP-91 and INT.HAP-91 was observed. On the 30th day, the defect was filled up completely in the same groups, result which was not observed in COL.HAP-91 group. Bone repair was complete in control group, HAP-91 group and INT.HAP-91 group (most premature), but in animals treated with COL.HAP-91 complete regeneration was not observed.

Polímero derivado de mamona acrescido de cálcio, associado ou não à medula óssea autógena na reparação de falhas ósseas

In order to evaluate tissue repair after the use of castor oil polymer implant additioned with 40% sodium carbonate, isolated or associated to autogenous bone marrow in heterotopic site and in experimental bone gaps in radii of rabbits, 30 White New Zealand rabbits were submitted to bilateral radial ostectomy. In 15 rabbits the bone gap of the right side was filled with polymer cylinders (group P) of similar size of the gaps; the remaining rabbits received autogenous bone marrow with the implant (group M). The bone defects of the left limb did not receive any treatment and served as control. Six rabbits received 6 implants in the Rectus abdominus muscle (heterotopic site) and in three of these rabbits the implants was associated with bone marrow. In the radiographic study both groups presented increased radiopacity at the implant site without bone axis deviation or resorption of the receptor bone ends. Group P presented irregular calcification areas at the peripheral region and over the polymer. Group M presented a more intense, regular and precocious radiopacity pattern in relation to group P. In microscopical evaluation there was evidence of immature bone tissue formation tending to organize itself, isolated sprouts of neoformed bone over the polymer and its superficial pores. It was concluded that implant allows osteogenenesis and osteoconduction in bone gaps, and bone formation was progressive, especially when additioned bone marrow aspirate; there was capillary, perivascular tissue and osteoprogenitor cells migrating into the pores; with fibrovascular tissue permeating implant surface; implants incorporation was slowly and was found incomplete until 9 weeks; the implant was able to induce foreign body reaction without toxic or secondary reactions to its presence. In heterotopic and orthotopic sites, the implant was not able to induce bone formation, inciting mild foreign-body reaction with giant cells and surrounding fibrous layer.

Avaliação histológica do processo de reparação da superfície articular de coelhos

The repairing process in the articular surface was evaluated microscopically in an osteochondral gap produced in the left troclear groove of 12 adult rabbits. It was observed that the granulation tissue wich initially filled this gap had diferentiated into hyaline cartilage by 4 weeks after surgery, presenting parcial reconstitution of subchondral bone excised at seven weeks following surgery. In some animals, cleft formation was observed between the repair tissue and adjacent bone, always where bone remodeling did not occured. In animals wich the bottom and lateral edges of the defect were formed by lamelar subchondral bone with few blood vessels, bone remodeling was not observed, having no attachment of the repair tissue to the adjacent bone. The results show the importance of complete removal of the lamelar subcondral bone during the surgical procedures in the articular surface, in order to allow better blood supply from the subjacent cancellous bone, better bone remodeling, and attachment of the repair tissue to the bottom of the defect.

Matriz óssea homóloga desmineralizada na preparação de falhas ósseas segmentares produzidas no rádio de coelhos

In order to evaluate osteoinduction and osteoconduction of the homologous demineralized bone matrix (DBM), processed in HCl 0.6 N solution for 24 hours and preserved in 70% ethanol solution, radial fragments were implanted in radius of adults rabbits. The animals were sacrificed at 3, 6, 9 and 12 post-operative weeks and the control group was sacrificed in 12 post-operative weeks. Radiographic exams of the operated radii were performed immediately after surgery and 3, 6, 9 and 12 post-operative weeks. After euthanasia, computed tomography, macroscopic, microscopic and tetracycline fluorescence exams were performed. DBM preserved its osteoinductive property and remains sterile. The empty spaces are favorable to osteoconduction and its growth factors induce osseous formation.

Potencial osteoindutor da matriz óssea homóloga desmineralizada de coelho

In order to evaluate osteoinduction of the homologous demineralized bone matrix (DBM), 1 cm radial fragments were demineralized in HCL 0.6 N solution for 24 hours, preserved in 70% ethanol solution and implanted in the abdominal musculature of 12 adult rabbits. The animals were sacrificed at 2, 4 and 6 post-operative weeks. After euthanasia, macroscopic, miroscopic and tetracycline fluorescence exams were performed. At four weeks, the implants already presented calcification signs. Microscopically, there was mature bone and medulary canal with hematopoietic cells, at six weeks. There was no evidence of rejection, or fluorescence at two and four weeks; at six weeks, there was difuse and concentric bands fluorescence. DBM in HCL 0.6 N solution and preserved in ethanol 70% solution maintains its osteoinductive property and remains sterile.

Caracterização de colágenos tipos I e III no estroma do carcinoma de células escamosas cutâneo em cães

O carcinoma de celulas escamosas (CCE) e uma neoplasia epitelial maligna que acomete caes e diversas outras especies, incluindo a humana. O CCE afeta varios sitios anatomicos e pode desenvolver metastase. O objetivo deste estudo foi a caracterizacao das fibras de colagenos tipos I e III no estroma do CCE cutâneo de caes. Para este trabalho, utilizaram-se 44 amostras de pele incluidas em parafina e que tiveram previo diagnostico de CCE. As amostras foram processadas histologicamente e coradas com hematoxilina/eosina para confirmacao do diagnostico e classificacao do grau de diferenciacao tumoral e com a coloracao histoquimica de picrosirius para observacao dos colagenos tipos I e III. O colageno tipo III mostrou maior expressao nos CCEs cutâneos bem diferenciados. O papel do colageno do tipo III nas neoplasias nao esta bem esclarecido, e outros fatores alem do grau de diferenciacao celular podem estar envolvidos em sua expressao e determinar sua importância na biologia tumoral.

Contribuição do plasma rico em plaquetas na reparação óssea de defeitos críticos criados em crânios de camundongos

No presente estudo, foram avaliados, de forma macro e microscopica, os resultados da aplicacao do PRP em defeitos osseos criticos de 6,0mm de diâmetro confeccionados em calvaria de 24 camundongos isogenicos C57BL/6 jovens, separados em dois grupos experimentais. O grupo controle nao recebeu tratamento, e no grupo tratado foram depositados, no interior do defeito, 50,0µL plasma em gel contendo 1,0x109 plaquetas. Constatou-se que o gel de PRP autologo depositado em defeitos criticos contribuiu positivamente para o processo de reparacao ossea, mormente na fase inicial.

Análise histomorfométrica da matriz extracelular do linfonodo poplíteo de cães naturalmente infectados por Leishmania (L.) chagasi

In the Americas, canine visceral leishmaniasis is caused by Leishmania (Leishmania) chagasi, an obligatory intracellular parasite of the phagocytic-monocytic system; the main histological changes associated with this disease occur in the lymphoid organs. Although dogs are considered to be the main carriers and disseminators of leishmaniasis in urban areas, there are few studies on the histopathologic and histomorphometric aspects in dogs naturally infected by L.chagasi analyzing the interaction between parasite and extracellular matrix. The current study characterize and quantify changes in the cellular and extracellular matrix (collagens type I and III) components of the popliteal lymph node from of 22 dogs with the natural infection by L. chagasi confirmed by indirect immuno-fluorescence assay (IFA) and compare theses findings with those fund in the popliteal lymph node from 10 non-infected dogs, that reacted negative in the IFA, and were clinically healthy. Lymph node fragments were longitudinally sliced and sections were processed for routine histopathology and stained by hematoxylin and eosin. For histomorphometry, additional sections from the same lymph node were fixed in glycol methacrylate and stained with toluidine blue. Lymph nodes from affected dogs were systemically enlarged, had increased numbers of lymphoid follicles, capsule hyperplasia and hypertrophy, and significant hyperplasia of lymphoid cells. In the lymph nodes from infected dogs, quantitative analyses of collagen fibers revealed predominance of type I collagen over type III fibers. These results demonstrate that dogs infected by L.chagasi experience degradation of the extracellular matrix components and consequently destruction of the lymphoid framework, thus altering nodal morphology.