Submitted by Kelson Anthony de Menezes (kelson@ucb.br) on 2016-12-19T19:37:39Z No. of bitstreams: 1 FabiodeBarrosAmaralDissertacao2014.pdf: 2938215 bytes, checksum: 3d899bf568eeae36d21979ba7aafab3c (MD5) / Made available in DSpace on 2016-12-19T19:37:39Z (GMT). No. of bitstreams: 1 FabiodeBarrosAmaralDissertacao2014.pdf: 2938215 bytes, checksum: 3d899bf568eeae36d21979ba7aafab3c (MD5) Previous issue date: 2014-03-31 / The equine chorionic gonadotropin (eCG) has been used as an important resource in the processes of reproduction in animals of economic interest, especially horses and cattle. The eCG molecule consists of a glycoprotein composed of two subunits called alpha and beta which together perform the function of both LH (luteinizing hormone) and FSH (follicle stimulating hormone) and due to this characteristic has great potential to induce ovulation in household species. Based on the aforementioned characteristics, the present work was was aimed at the study and characterization of the structure of eCG using molecular modeling techniques as well as testing its production in cells of the microorganism Pichia pastoris. Furthermore, chimeric models have been proposed linking the eCG and immunoglobulin G (IgG) molecule based on studies that reveal the great potential of it in raising the half-life of other molecules circulating in the organism. Four chimeric models were prepared so that the eCG alpha and beta chains are connected by a hinge, short and flexible regions, while the terminal region of the alpha chain was connected to the N-terminal portion of the CH2 domain of the IgG by another hinge region, thus resulting in a model in which two different molecules are linked. / A gonadotrofina cori??nica equina (eCG) tem sido um importante recurso usado nos processos de reprodu????o em animais de interesse econ??mico, principalmente equinos e bovinos. A mol??cula de eCG consiste em uma glicoprote??na composta de duas subunidades denominadas alfa e beta que juntas desempenham a fun????o tanto de LH (horm??nio luteinizante) como FSH (horm??nio fol??culo estimulante) e devido a essa caracter??stica possui um grande potencial para induzir a ovula????o em esp??cies dom??sticas. Baseando-se nas caracter??sticas citadas, o presente trabalho teve como objetivo estudar e caracterizar a estrutura do eCG utilizando-se t??cnicas de modelagem molecular assim como testar a sua produ????o em c??lulas do microrganismo Pichia pastoris. Al??m disso, foram propostos modelos quim??ricos ligando eCG ?? mol??cula de imunoglobulina G (IgG) tendo como base estudos que revelam o grande potencial da mesma em elevar a meia vida de outras mol??culas em circula????o no organismo. Quatro modelos quim??ricos foram apresentados de forma que as cadeias beta e alfa do eCG s??o conectadas por regi??es hinge que s??o sequ??ncias curtas e flex??veis da mol??cula enquanto que a regi??o terminal da cadeia alfa foi conectada por meio de outra regi??o hinge, ?? por????o N-terminal do dom??nio CH2 da IgG, portanto, resultando em um modelo em que est??o ligadas duas mol??culas diferentes.

Submitted by Setor de Tratamento da Informa??o - BC/PUCRS (tede2@pucrs.br) on 2015-06-24T12:10:10Z No. of bitstreams: 1 471057 - Texto Completo.pdf: 1136786 bytes, checksum: 1d97578e696595fa93bee351f70a6852 (MD5) / Made available in DSpace on 2015-06-24T12:10:10Z (GMT). No. of bitstreams: 1 471057 - Texto Completo.pdf: 1136786 bytes, checksum: 1d97578e696595fa93bee351f70a6852 (MD5) Previous issue date: 2015-03-31 / Growth factors such as the recombinant human growth hormone are drawing attention on the field of bone reconstruction. However, due to the protein?s short half-life, biodegradable polymers, like PLGA, have been applied seeking a slower and controlled drug release for topical applications. Objective: To asses, in vitro, the behavior of PLGA matrices with 2 different rhGH concentrations. Materials and Methods: Three groups of triplicate samples were created: group I with pure PLGA (control), group II with PLGA loaded with a higher rhGH concentration and group III with PLGA loaded with a lower concentration of the hormone. Each group was analyzed macro and microscopically (SEM) and, by means of the hidrolitic degradation test, the dynamics of release of rhGH as well as the alterations on the pH of the incubation medium were evaluated, throughout 29 days. Results: Matrices from all groups showed indistinguishable macro and microscopic features. Group II released a higher quantity of rhGH, as expected, however there was no clear release pattern on both groups. rhGH release was observed until the 22? day and there was no statistically significant difference on weight loss between the 3 groups. pH values found were clinically viable. Conclusion: Morphological properties of the matrices must be assessed according to its applications and target-tissue to be substituted/regenerated and it was not possible to determine a pattern of rhGH distribution within PLGA matrices on this survey. The matrices can be used as rhGH carriers and were able to extend hormone release for a period of 22 days. Wheight loss showed equal patterns (statistically), despite the addition of rhGH to the polymer. The pH values were clinically viable and the reduction of rhGH concentrations on future researches should not be carried out using this methodology, possibly owing to limitations on screening with ultra-violet spectroscopy. / Fatores de crescimento como o horm?nio do crescimento humano recombinante v?m ganhando aten??o no campo da reconstru??o ?ssea. No entanto, devido ? sua meia-vida curta, pol?meros biodegrad?veis, como o PLGA, v?m sendo utilizados visando libera??o mais lenta e controlada deste horm?nio quando de sua aplica??o t?pica. Objetivo: Avaliar, in vitro, o comportamento de matrizes de PLGA com 2 concentra??es diferentes de rhGH. Materiais e M?todos: Tr?s grupos de amostras em triplicatas foram montados: grupo I com PLGA puro (controle); grupo II com PLGA associado ao rhGH em maior concentra??o e; grupo III com PLGA associado ao rhGH em menor concentra??o. Cada grupo foi analisado macroscopicamente e microscopicamente (MEV) e, por meio de teste de degrada??o hidrol?tica, avaliou-se a din?mica de libera??o do rhGH assim como as altera??es de pH no meio de incuba??o ao longo de 29 dias. Resultados: As matrizes dos 3 grupos de estudo ficaram com caracter?sticas morfol?gicas indistingu?veis macro e microscopicamente. O grupo II liberou maior quantidade de rhGH que o grupo III, como era o esperado, no entanto, n?o houve um padr?o claro de libera??o em ambos os grupos. A libera??o de rhGH foi observada at? o 22? dia de avalia??o e n?o houve diferen?a estat?stica na perda de massas entre os grupos. Os valores de pH encontrados foram compat?veis com a aplica??o cl?nica. Conclus?o: As propriedades morfol?gicas devem ser avaliadas conforme a aplica??o do material e do tecido alvo a ser regenerado/substitu?do e n?o foi poss?vel estabelecer um padr?o de distribui??o do rhGH nas matrizes de PLGA neste estudo. As matrizes podem ser utilizadas como carreadoras de rhGH tendo sido capazes de prolongar a libera??o do horm?nio por um per?odo de 22 dias. A perda de massas se manteve igual (estatisticamente), a despeito da adi??o do rhGH ao pol?mero. O pH das amostras se mostrou favor?vel ? aplica??o cl?nica das matrizes e a redu??o nas concentra??es de rhGH para estudos futuros n?o deve ser avaliada por esta metodologia, visto que podem se tornar de dif?cil leitura pelo EAU-Vis.

Submitted by Caroline Xavier (caroline.xavier@pucrs.br) on 2017-05-25T15:00:50Z No. of bitstreams: 1 DIS_RICARDO_GIACOMINI_DE_MARCO_PARCIAL.pdf: 425010 bytes, checksum: 70ab63ee01f40dabcc01f10861fb479f (MD5) / Made available in DSpace on 2017-05-25T15:00:50Z (GMT). No. of bitstreams: 1 DIS_RICARDO_GIACOMINI_DE_MARCO_PARCIAL.pdf: 425010 bytes, checksum: 70ab63ee01f40dabcc01f10861fb479f (MD5) Previous issue date: 2017-03-06 / At the back regions of maxilla, often are unable to accommodate dental implants because of thin bone thickness. In these cases, the procedure of lifting the maxillary sinus floor is the viable option to rehabilitate this area. One of the main reasons patients give up treatment is the waiting time between the bone graft and the installation of dental implants (around 6 months). This research sought to use Human Growth Hormone (GH), together with Lyophilized Bovine Bone (Bio-Oss?), in Maxillary Sinus Floor Survey procedures to analyze whether there is a reduction in bone maturation time. In the first phase of the work, three patients were operate bilaterally, and on one side the conventional maxillary sinus survey and on the other side was add Human Growth Hormone (Somatropin). After 3 to 5 months were make Computed tomography of cases operated. The images obtained were assemble in questionnaire format, without identification of the patient, material used, time after surgery and use of GH or not. The questionnaire was forwad to 52 Oral and Maxillofacial Surgeons, by e-mail. At the end of the work, we concluded that growth hormone-enhanced grafts obtained full majority of the evaluators' agreement to install the implants at the time the Topographies were make. Unlike in the group without Growth Hormone, the great majority of the evaluators considered the moment inopportune for implant installation. These results demonstrate positive interference in the quality of the tomographic image, when the increase of the Growth Hormone was make, according to the opinion of the evaluators. Key / Regi?es posteriores de maxila, frequentemente, s?o incapazes de acomodar implantes dent?rios devido a pouca espessura de osso. Nesses casos, o procedimento de levantamento do assoalho do seio maxilar ? a op??o vi?vel para reabilitar essa ?rea. Um dos principais motivos que faz os pacientes desistirem do tratamento ? o tempo de espera entre o enxerto ?sseo e a instala??o dos implantes dent?rios (em torno de 6 meses). Esta pesquisa buscou utilizar o Horm?nio de Crescimento Humano Recombinante (rhGH), juntamente com o Osso Bovino Liofilizado (Bio-Oss?), em procedimentos de Levantamento de Assoalho de Seio Maxilar para analisarmos se h? redu??o no tempo de matura??o ?ssea. Na primeira fase do trabalho, tr?s pacientes foram operados bilateralmente, sendo que, em um lado, foi realizado o levantamento de seio maxilar convencional e, no outro lado, foi acrescentado o rhGH (Somatropina). Entre tr?s a cinco meses, foram realizadas tomografias computadorizada dos casos operados. As imagens obtidas foram montadas em formato de question?rio, sem identifica??o de paciente, material utilizado, tempo p?s-operat?rio e uso ou n?o do rhGH. O question?rio foi encaminhado para cinquenta e dois Cirurgi?es Bucomaxilofaciais e Implantodontistas por e-mail. Ao final do trabalho, conclu?mos que os enxertos com incremento do rhGH obtiveram plena maioria da concord?ncia dos avaliadores em instalar os implantes no momento em que as tomografias foram realizadas. Ao contr?rio, no grupo sem incremento do rhGH, a grande maioria dos avaliadores considerou o momento inoportuno para instala??o dos implantes. Esses resultados demonstram interfer?ncia positiva na qualidade da imagem tomogr?fica, quando realizado o incremento do Horm?nio de Crescimento, conforme a opini?o dos avaliadores.

Submitted by Caroline Xavier (caroline.xavier@pucrs.br) on 2017-06-26T14:56:31Z No. of bitstreams: 1 TES_RICARDO_FERNANDES_GARCIA_PARCIAL.pdf: 159331 bytes, checksum: aacf8f2e7202562d6ba8bfb886ab0c8d (MD5) / Made available in DSpace on 2017-06-26T14:56:31Z (GMT). No. of bitstreams: 1 TES_RICARDO_FERNANDES_GARCIA_PARCIAL.pdf: 159331 bytes, checksum: aacf8f2e7202562d6ba8bfb886ab0c8d (MD5) Previous issue date: 2017-03-08 / More and more studies are focusing on the combination of matrices that have osteoconductive characteristics with osteoinductive proteins. These proteins can stimulate the differentiation of mesenchymal and osteoprogenitor cells into osteoblasts and thereby increase the migration of cells related to bone formation within the defect site. The main materials used for these purposes are biodegradable polymers, such as PLA (poly lactic acid) and PLGA (poly lactic glycolic acid). Several drugs are used to be released in these systems, such as antibiotics, contraceptives and proteins, including human growth hormone (GH). RhGH in bone tissue promotes the increased deposition of proteins by chondrocytes and osteoblasts, an increase in the number of mitoses and the conversion of chondrocytes into osteoblasts. Thus, it is necessary to evaluate this biomaterial (PLGA \ rhGH) as a controlled release device. Thus achieving the presence of rhGH in the site of bone healing. For this study was used as animal model, wistar rats. The study was carried out in accordance with Law No. 11,794 of October 8, 2008, as well as following the Brazilian Directive of Practice for the Care and Use of Animals for Scientific and Educational Purposes - DBPA of CONCEA. Project approved by CEUA of PUCRS under number 15/00461. Thirty adult wistar rats were used, in which they were submitted to bone defects with Carbide spherical drill number 704, the defect corresponded to the diameter of the drill. The rats were submitted to the same treatment but suffered euthanasia at different times (07, 15 and 20 days). The femurs of the rats were radiographed in the range of 7, 15, and 20 days to gauge the optical density. Slides were then produced for histological analysis, with cuts of approximately 5 ?m thick and stained with hematoxylin / eosin (HE). Three more representative cuts of each blade were chosen. The systemic repercussion of rhgh was assessed through IGF-1 levels through blood collection that was performed before each euthanasia, in the control group and in the experimental group. The blood collected was centrifuged (2500 revolutions per minute for 10 min) and serum obtained stored in a freezer at -20 ? C for further determination of IGF-1 plasma levels using the method based on an enzymatically amplified assay of the " Sandwich, "held at the Senhor dos Passos laboratory in Porto Alegre, RS. For statistical analysis, the Tukey test was applied, 5% significance level and ANOVA variance analysis. There were no statistically significant differences in bone mineral densities, however numerically the PLGA\RhGH grafts always had a greater amount of shades of gray. In conclusion, defects with PLGA\RhGH grafts showed a higher optical density. A higher mineral density compared to the autogenous graft In the histological analyzes there was also no statistically significant difference between the autogenous and PLGA\RhGH grafts. However, by numerical analysis we observed a better and equal performance between the PLGA\RhGH graft and autogenous graft. Defects with PLGA\RhGH grafting probably had faster healing and maturation than autogenous grafts. / Cada vez mais estudos est?o focando a combina??o de matrizes que possuam caracter?sticas osteocondutora com prote?nas osteoindutivas. Essas prote?nas podem estimular a diferencia??o de c?lulas mesenquimais e osteoprogenitoras em osteoblastos e assim aumentar a migra??o de c?lulas relacionadas ? forma??o ?ssea dentro do s?tio do defeito. Os principais materiais utilizados para esses objetivos s?o os pol?meros biodegrad?veis, como o PLA (poli ?cido l?tico) e o PLGA (poli ?cido glic?lico l?tico). Diversas drogas s?o utilizadas para serem liberadas nesses sistemas, como antibi?ticos, anticoncepcionais e prote?nas, incluindo o horm?nio do crescimento humano recombinante (rhGH). O rhGH no tecido ?sseo, promove a deposi??o aumentada de prote?nas pelos condr?citos e osteoblastos, aumento do n?mero de mitoses e a convers?o de condr?citos em osteoblastos. Desta forma se faz necess?rio a avalia??o desse biomaterial (PLGA\rhGH) como dispositivo de libera??o controlada. Conseguindo com isso a presen?a do rhGH intimamente no local da cicatriza??o ?ssea. Para este estudo foi utilizado como modelo animal,ratos wistar. O estudo foi todo realizado de acordo com a Lei N0 11.794, de 8 de outubro de 2008 bem como seguindo a diretriz Brasileira de Pr?tica para o Cuidado e a Utiliza??o de Animais para Fins Cient?ficos e Did?ticos ? DBPA do CONCEA. Projeto aprovado pelo CEUA da PUCRS sob n?mero 15/00461. Foram utilizados trinta ratos wistar adultos, nos quais foram submetidos a defeitos ?sseos nos f?mures com broca, os defeitos tinham todos mais ou menos 05mm de di?metro. Os ratos foram submetidos ao mesmo tratamento, por?m divididos em tr?s grupos de acordo com o tempo das eutan?sia :Grupo 01 eutan?sia em 07dias , Grupo 02 eutan?sia em 15 dias e Grupo 03 eutan?sia em 20 dias. Os f?mures dos ratos foram radiografados no intervalo entre 07, 15, e 20 dias para aferir a densidade ?ptica. Em seguida foram produzidas l?minas para an?lises histol?gicas, com cortes de aproximadamente 5?m de espessura e corados com hematoxilina/eosina (HE). Foram escolhidas tr?s cortes mais representativos de cada l?mina. A repercuss?o sist?mica do rhGH foi avaliada atrav?s dos n?veis de IGFI atrav?s da coleta de sangue que foi realizada antes de cada eutan?sia, no grupo-controle e no grupo experimental. O sangue coletado foi centrifugado (2500 rota??es por minuto por 10 min) e o soro obtido armazenado em freezer a -20?C para posterior determina??o dos n?veis plasm?ticos de IGFI usando o m?todo que se baseia em um ensaio enzimaticamente amplificado do tipo ?sandu?che?, realizado no laborat?rio Senhor dos Passos em Porto Alegre,RS. Para an?lise estat?stica foi aplicado o Teste Tukey, n?vel de 5% de signific?ncia e an?lise de vari?ncia ANOVA. N?o houve diferen?a estatisticamente significante em rela??o as densidades minerais ?sseas, por?m numericamente os enxertos com PLGA\rhGH tiveram sempre uma quantidade maior de tons de cinza. Conclui-se que os defeitos com enxerto de PLGA\rhGH apresentaram uma maior densidade ?ptica e uma maior densidade mineral em compara??o ao enxerto aut?geno. Nas an?lises histol?gicas tamb?m n?o houve diferen?a estatisticamente significante entre os enxertos aut?geno e de PLGA\rhGH. Por?m pelas analises num?ricas observamos um desempenho melhor e/ou igual entre o enxerto de PLGA\rhGH e enxerto aut?geno. Os defeitos com enxerto de PLGA\rhGH, de ocordo com a metodologia empregada, densidade mineral ?ptica e pelas l?minas histol?gicas, observamos uma cicatriza??o e matura??o mais r?pida do que os enxerto aut?genos.

Submitted by Setor de Tratamento da Informa??o - BC/PUCRS (tede2@pucrs.br) on 2015-05-14T20:32:00Z No. of bitstreams: 1 468626 - Texto Completo.pdf: 36893570 bytes, checksum: f40ac0cf663ae01a836d6017460f63fc (MD5) / Made available in DSpace on 2015-05-14T20:32:00Z (GMT). No. of bitstreams: 1 468626 - Texto Completo.pdf: 36893570 bytes, checksum: f40ac0cf663ae01a836d6017460f63fc (MD5) Previous issue date: 2015-03-27 / Objective: to evaluate the stability of PLGA 50:50 formulations incorporated with recombinant human growth hormone (rhGH) under different storage time and temperatures. Material and Methods: a total of 27 macrospheres of PLGA + rhGH were prepared using 7.5g of PLGA and 1,33mg of rhGH. Twenty-four of the 27 samples were equally divided into 4 groups (I, II, III and IV), according to the storage time, and two subgroups (A and F), according to the storage temperature prior to degradation. The remaining 3 samples were degraded immediately after its preparation (group 0). The concentration of rhGH released from the matrices was measured by UV-visible spectrophotometry absorption analysis. A calibration curve defined rhGH concentration values based on the final absorbance values. All colected data were submitted to descriptive and analytical statistical analysis with a confidence interval of 95% (p <0.05). Results: in general, the samples stored for different periods of time followed similar rhGH release rate of the samples belonged to group 0, with peaks of major and minor release rates and showing an important release until the 14th day of degradation, but no statistically significant differences between the study groups. Although higher absorbance values were constantly found for the matrices in subgroup F, the pattern of rhGH release was similar and no statistically significant difference was evident between the subgroups compared. The pH analysis revealed a lower pH for the matrices stored refrigerated, with no statistical difference. Conclusion: PLGA + rhGH formulations are stable. Until the 28th day, storage time and temperature do not affect the stability of the formulation. / Objetivo: avaliar a estabilidade das formula??es de PLGA 50:50 incorporadas com horm?nio de crescimento recombinante humano (rhGH) sob diferentes per?odos e temperaturas de armazenamento. Materiais e m?todos: um total 27 matrizes de PLGA + rhGH foram confeccionadas utilizando 7,5g de PLGA e e 1,33mg de rhGH. Vinte e quatro das 27 matrizes foram divididas igualmente em 4 grupos (I, II, III e IV), de acordo com o tempo de armazenamento, e em 2 subgrupos (A e F), de acordo com a temperatura de armazenamento previamente ? sua degrada??o. As 3 matrizes restantes foram degradas imediatamente ap?s sua confec??o (grupo 0). A concentra??o de rhGH liberado das matrizes foi mensurada em dias determinados atrav?s da an?lise de espectrofotometria de absor??o UV-vis?vel do sobrenadante removido do tubos de ensaio onde as matrizes foram degradadas. Uma curva de calibra??o definiu os valores de concentra??o de rhGH baseado nos valores finais de absorb?ncia. Os dados obtidos foram submetidos ? an?lises estat?sticas descritivas e anal?ticas com intervalo de confian?a de 95% (p<0,05). Resultados: em geral, as matrizes armazenadas por diferentes per?odos de tempo seguiram o mesmo padr?o de libera??o de rhGH das matrizes do grupo 0, com picos de maior e menor libera??o entre os dias avaliados e demonstrando uma libera??o expressiva at? o 14o dia de degrada??o, mas sem diferen?as estatisticamente significativas entre grupos do estudo. Apesar de constantemente haver maiores valores de absorb?ncia para as matrizes do subgrupo F, o padr?o de libera??o de rhGH foi similar para os dois subgrupos, e n?o houve diferen?a estatisticamente significante entre os dois subgrupos comparados. A an?lise de pH revelou menor pH para as matrizes armazenadas em ambiente refrigerado, sem diferen?a estat?stica. Conclus?o: as formula??es de PLGA incorporadas com rhGH s?o est?veis. At? 28 dias, o tempo e a temperatura de armazenamento n?o influenciam na estabilidade da formula??o.

Made available in DSpace on 2015-04-14T13:30:16Z (GMT). No. of bitstreams: 1 443201.pdf: 4831267 bytes, checksum: 8dd21e93476eb996bc1ebeb289330d2b (MD5) Previous issue date: 2012-08-31 / Osteoarthritis is a slowly progressive degenerative disease affecting several joints of the human body, including the TMJ. Tissue engineering has advanced to reduce osteochondral defects and to regenerate the TMJ. Nineteen rabbits were used in the present study. Eighteen of them had the articular discs of the TMJ removed in order to induce osteoarthritis. The animals were divided into three groups of six rabbits: group A - five rabbits treated with injections of rhGH (0.5 IU/kg) in the right joint and NaCl (0.9%) in the left joint in 14 days; group B - five rabbits received injections of rhGH (0.5 IU/kg) in the right joint and NaCl (0.9%) in the left joint in 14 and 21 days; and group C - five rabbits underwent injections of rhGH (0.5 IU/kg) in the right joint and NaCl (0.9%) in the left joint in 14, 21, and 28 days. The sixth rabbit of each group was the control and did not receive injections. The 19th rabbit was used as a normal TMJ control and did not undergo surgery. The animals were evaluated using cone beam computed tomography and histological examination. CT showed that the joints treated with three injections of 0.9% NaCl and the control group had higher rates of osteoarthritis (p < 0.05). Histological analysis revealed that the joints treated with rhGH had higher tissue cellularity and thickness in group A, whereas the joints treated with rhGH and NaCl (0.9%) had high tissue cellularity and thickness in group B than in the control group. There was no statistical difference in terms of degree of severity of osteoarthritis. / A osteoartrite ? uma doen?a lentamente progressiva e degenerativa que atinge v?rias articula??es do corpo humano, incluindo a ATM. A engenharia tecidual tem avan?ado no sentido de reduzir os defeitos osteocondrais e regenerar essa articula??o. Foram utilizados no estudo 19 coelhos, dos quais 18 tiveram os discos articulares da ATM removidos, com o intuito de induzir osteoartrite. Os animais foram divididos em 3 grupos com 6 cobaias: grupo A, onde 5 coelhos realizaram 1 infiltra??o na articula??o direita com rhGH (0,5UI/kg) e na esquerda com NaCl (0,9%) em 14 dias; grupo B, onde 5 coelhos realizaram 1 infiltra??o na articula??o direita com rhGH (0,5UI/kg) e na esquerda com NaCl (0,9%) em 14 e 21 dias; grupo C, onde 5 coelhos realizaram 1 infiltra??o na articula??o direita com rhGH (0,5UI/kg) e na esquerda com NaCl (0,9%) em 14, 21 e 28 dias. O sexto coelho de cada grupo n?o recebeu infiltra??es e serviu como controle. O 19? coelho n?o foi operado e serviu como controle de ATM normal. Os animais foram avaliados por tomografia computadorizada cone beam e histologicamente. Os resultados demonstraram que, tomograficamente, as articula??es tratadas com 3 infiltra??es de NaCl 0,9% e o grupo controle obtiveram maiores ?ndices de osteoartrite (p<0,05). Na an?lise histol?gica, as articula??es tratadas com rhGH apresentaram maior celularidade e espessura tecidual no grupo A, no grupo B, as articula??es infiltradas com rhGH e NaCl (0,9%) apresentaram maior celularidade e espessura que no grupo controle. Quanto ao grau de severidade da osteoartrite, n?o houve diferen?a estat?stica.

Made available in DSpace on 2015-04-14T13:30:23Z (GMT). No. of bitstreams: 1 448352.pdf: 2234936 bytes, checksum: 8f2dcd05e3619f61d684fbf373e529c4 (MD5) Previous issue date: 2013-03-15 / Objective: To incorporate recombinant human growth hormone in a biodegradable polymer (PLGA) Material and Methods: It was used a blend of two solvents, methanol and dichloromethane in a ratio by weight of 1.5: 5.0 for dissolving the PLGA. Another solution was made with rhGH, zinc acetate and deionized water. The two mixtures were then mixed together under mechanical stirring and poured into silicone molds circular of 01cm in diameter and around 02mm of espessur. This mixture was left in the evaporation chamber of solvent for 48 hours. The matrices were then degraded in vitro in PBS pH 7.4, thermostatted bath. The samples were removed from the bath in the range of 01, 02, 03, 04, 07, 10, 14 days. Was measured weight loss, pH and concentration of hormone released versus time. Results: The concentration of hormone released versus time was increased until the third day. On the fourth day had a fall and on the seventh day there have been increased hormone released by the tenth day, the fourteenth day was falling again. The pH had a sharp drop from 7.4 to 3.2 on the first day and keeping a small drop until the fourteenth day. The mass loss was a gradual loss in relation to time as was to be expected. Conclusion: The PLGA is a good biomaterial for making this type of device. You can incorporate the rhGH this array to get a possible material that may serve as a bone graft. / Objetivo: Incorporar o horm?nio de crescimento recombinante humano em um pol?mero biodegrad?vel o poli ?cido l?ctico glic?lico ( PLGA ) para utiliza??o na ?rea de cirurgia e traumatologia bucomaxilofacial. Materiais e M?todos: Foi utilizada mistura de dois solventes, metanol e o diclorometano numa propor??o em massa de 1,5 : 5,0 para dissolver o PLGA. Foi feita outra solu??o com o rhGH (horm?nio de crescimento humano recombinante), acetato de zinco e ?gua deionizada. As duas misturas foram ent?o misturadas sob agita??o mec?nica e vertidas em moldes de silicone circular de 01cm de di?metro e aproximadamente 02mm de espessura. Essa mistura foi deixada na c?mara de evapora??o de solvente por 48 horas. As matrizes, ent?o, foram degradadas in vitro em PBS (solu??o tamp?o fosfato salino) com pH 7.4, em banho termostatizado. As amostras foram retiradas do banho no intervalo de 01, 02, 03, 04, 07, 10, 14 dias. Foi aferida a perda de massa, varia??o do pH e concentra??o do horm?nio liberado em fun??o do tempo. Resultados: A concentra??o do horm?nio liberado em fun??o do tempo foi aumentando ate o terceiro dia. No quarto dia, houve uma queda e, no s?timo, ocorreu um aumento do horm?nio liberado, estendendo-se at? o d?cimo dia, no d?cimo quarto dia houve queda novamente. O pH teve uma queda brusca de 7.4 para 3.2 no primeiro dia e mantendo uma pequena queda at? o d?cimo quarto dia. A perda de massa foi gradual, em rela??o ao tempo, como j? era esperado. Conclus?o: O PLGA ? um bom biomaterial para esses fins. Revelou-se poss?vel incorporar o rhGH nessa matriz, de modo a, ent?o, desenvolver-se um poss?vel material que sirva como enxerto em tecido ?sseo.

Made available in DSpace on 2015-04-14T13:30:36Z (GMT). No. of bitstreams: 1 459060.pdf: 1405009 bytes, checksum: 550db90a424df77982f137b221a18c73 (MD5) Previous issue date: 2014-02-24 / The PLGA is a bioabsorbable polymer derived from lactic and glycolic acids which has been used as drug release system . One of the drugs with the potential to use this type of vehicle is the recombinant human growth hormone (rhGH), because it has advantages such as speed up the bone integration process , bone maturation and remodeling around implants, in addition to stimulating osteogenesis. Objective: This study aims to evaluate the matrix degradation of PLGA and rhGH. Materials and Methods: 75:25 (Group I) and 50:50 (Group II): Two ratios of PLGA were used. The matrices were prepared by solvent evaporation technique as it follows: firstly, the PLGA was dissolved in dichloromethane and methanol and then it was mixed with a composed mixture of recombinant human growth hormone, zinc acetate, solution and deionized water. This mixture was poured into silicone circular shape moulds approximately 10 mm diameter which were placed in a solvent evaporation chamber at room temperature, where they were dried by nitrogen gas force action. The control samples were made just dissolving PLGA and pouring it into the silicone moulds. After the drying period, all the samples were weighed on a precision scale. Then, they were immersed in buffer solution and the hydrolytic degradation experiment was investigated and the samples remained for a predetermined period of time. After this process was finished, the solid sample portion was dried and weighed again. The supernatant went through pH analysis and later, by UV-visible absortion spectrophotometry analysis. Results:It was possible to calculate the concentration of rhGH released in the middle through the straight line obtained in a calibration curve.We observed that, from the first to the last day the released rhGH concentration in the medium was higher in Group II. The loss of mass percentage increased over the time in both groups. But this loss of mass was higher in group I compared to Group II. The pH showed a little variation over the time. The average was between 7.5 and 7.9. Conclusion:The matrices which were prepared with PLGA 50:50 showed the best results in relation to the ones prepared with PLGA 75:75.These are some advantages: easy handling, more uniform pores, less bubbles and the growth hormone was released slowly and gradually. / O PLGA ? um pol?mero bioabsorv?vel derivado dos ?cidos l?ctico e glic?lico que tem sido usado como sistema de libera??o de drogas. Uma das drogas com potencial de utiliza??o nesse tipo de ve?culo ? o horm?nio do crescimento humano recombinante (rhGH), pois apresenta vantagens como, por exemplo, acelerar o processo de osteointegra??o, remodela??o ?ssea e matura??o ao redor de implantes, al?m de estimular a osteog?nese. Objetivo: Este trabalho tem por objetivo avaliar a degrada??o de matrizes de PLGA com rhGH. Materiais e M?todos: Foram utilizadas duas propor??es de PLGA: 75:25 (Grupo I) e 50:50 (Grupo II). As matrizes foram confeccionadas por meio da t?cnica de evapora??o de solventes da seguinte forma: primeiramente o PLGA foi dissolvido com diclorometano e metanol e, posteriormente, foi misturado ? uma solu??o composta por horm?nio do crescimento humano recombinante, acetato de zinco e ?gua deionizada. A mistura dessas subst?ncias foi vertida em moldes de silicone de formato circular, com aproximadamente 10 mm de di?metro, os quais foram acondicionados em uma c?mara de evapora??o de solvente, ? temperatura ambiente, onde as matrizes foram secas por meio do arraste for?ado pela a??o de g?s nitrog?nio. As amostras controle, entretanto, foram confeccionadas apenas dissolvendo o PLGA e vertendo-o nos moldes de silicone. Ap?s o per?odo de secagem todas as amostras foram pesadas em uma balan?a de precis?o. Na sequ?ncia, as matrizes foram imersas em solu??o tamp?o e realizou-se o ensaio de degrada??o hidrol?tica, no qual as amostras permaneceram por um per?odo de tempo pr?-determinado. Terminado esse processo, a parte s?lida da amostra foi seca e, novamente, pesada. O sobrenadante, ent?o, passou pela an?lise de pH e pela an?lise de espectrofotometria de absor??o UV-vis?vel. Resultados: Atrav?s da equa??o da reta obtida em uma curva de calibra??o, foi poss?vel calcular a concentra??o do rhGH liberado no meio. Com isso, observou-se que, do primeiro at? o ?ltimo dia a concentra??o de rhGH liberado no meio foi maior no Grupo II. O percentual de perda de massa foi crescente ao longo tempo, em ambos os grupos. Por?m essa perda de massa foi maior no Grupo I, em rela??o ao Grupo II. O pH apresentou pouca varia??o ao longo do tempo. A m?dia ficou entre 7,5 e 7,9. Conclus?o: As matrizes preparadas com PLGA na propor??o 50:50 apresentaram melhores resultados em rela??o ?s matrizes preparadas com PLGA 75:25. Dentre as vantagens, ressaltam-se: maior facilidade de manipula??o, poros mais uniformes, menos bolhas de ar e libera??o do horm?nio do crescimento de forma mais lenta e gradual.

Submitted by Setor de Tratamento da Informa??o - BC/PUCRS (tede2@pucrs.br) on 2016-05-17T20:18:03Z No. of bitstreams: 1 TES_ALEXANDRE_DA_SILVEIRA_GERZSON_COMPLETO.pdf: 8833517 bytes, checksum: b4672493ffabfc3e5c284282aa067e14 (MD5) / Made available in DSpace on 2016-05-17T20:18:03Z (GMT). No. of bitstreams: 1 TES_ALEXANDRE_DA_SILVEIRA_GERZSON_COMPLETO.pdf: 8833517 bytes, checksum: b4672493ffabfc3e5c284282aa067e14 (MD5) Previous issue date: 2016-03-23 / Biomaterials, as an alternative to autogenous bone and other biological tissues, have been widely used in oral and maxillofacial surgery. In this context, a biomaterial that functions as a scaffold (osteoconductor), combined with a growth factor (osteoinductor), would be of great interest for clinical application. Biodegradable polymers used for slow drug release have been investigated, demonstrating good results and interesting potential. Growth hormone (rhGH) may be released by incorporating it into these polymers. This study aimed to evaluate cell adhesion and proliferation of a polymeric biomaterial for slow release of rhGH. PLGA and PLGA/PCL (at a 70/30 ratio of PLGA to PCL) matrices were prepared by the solvent evaporation method, combined or not with GH. The biomaterials were tested for toxicity and cell viability using an MTT assay with NIH3T3 mouse cells (ATCC). Cell toxicity was assessed at 24, 48, 72 hours, and 7 days of biomaterial exposure to culture medium. After were tested?for cell adhesion and proliferation by culture in mesenchymal stem cells derived from Wistar rat bone marrow, DAPI staining, and subsequent cell counting, in addition to scanning electron microscopy. Cell adhesion and proliferation was assessed at 24 and 72 hours of biomaterial exposure to culture medium. All polymers had high cell viability rates. However, from 48 hours onwards, the groups with rhGH-polymer combinations had better results than the polymer groups without association with GH when compared to the control group. At 7 days of culture, only the pure PLGA matrix showed a significant difference from the control group. These results may suggest a preference of cells for the presence of rhGH in the biomaterial in culture medium? especially in the PLGA matrix. GH appeared to contribute to the increase in cell viability observed at some assessment time points, especially when combined with PLGA as compared to pure PLGA. All tested polymers exhibited cell adhesion and proliferation. However, PLGA-based biomaterials, especially when combined with rhGH, showed greater cell proliferation when the difference in growth from 24 to 72 hours was evaluated. rhGH appeared to modify the polymer surface, with increased roughness and microporosity. This feature was more evident in the PLGA+rhGH combination. Further studies are required to clarify this potential for development of new biomaterials. / Biomateriais como alternativas ao osso aut?geno e outros tecidos biol?gicos, s?o muito utilizados no tratamento de pacientes na rotina cir?rgica da regi?o maxilofacial. Neste contexto, um biomaterial com caracter?sticas de arcabou?o (osteocondutor), e associado ? um fator de crescimento (osteoindutor), seria de grande interesse para pesquisa e aplica??o cl?nica. Pol?meros biodegrad?veis, utilizados para a libera??o lenta de medicamentos, vem sendo estudados, demonstrando bons resultados e interessante potencial. o rhGH pode ser liberado atrav?s da sua incorpora??o ? estes pol?meros. Desta forma, se faz necess?rio a avalia??o da biocompatibilidade e toxicidade de um biomaterial polim?rico para libera??o lenta do medicamento. Atrav?s da t?cnica de evapora??o de solvente foram preparados matrizes de PLGA e PLGA/PCL na raz?o 70/30 de PLGA e PCL, respectivamente, associados ou n?o ao rhGH. Os biomateriais foram testados relacionados ? sua toxicidade e viabilidade celular, atrav?s de um ensaio de MTT com c?lulas de camundongo NIH 3T3 ATCC. A avalia??o da toxicidade celular foi realizada nos tempos de 24 horas, 48 horas, 72 horas e 7 dias de exposi??o dos biomateriais com o meio de cultura. Ap?s, os biomateriais foram testados relacionados ? sua ades?o e prolifera??o celular, atrav?s do cultivo em c?lulas-tronco mesenquimais de rato Wistar, com ensaio de colora??o DAPI e posterior contagem celular, al?m de microscopia eletr?nica de varredura. A avalia??o da ades?o e prolifera??o celular foi realizada nos tempos de 24 e 72 horas de exposi??o dos biomateriais com o meio de cultura. Todos os pol?meros testados apresentaram altas taxas de viabilidade celular, por?m, os grupos associados ao rhGH parecem demonstrar melhores resultados do que os grupos de pol?meros sem associa??o ao horm?nio quando comparados ao grupo controle em alguns per?odos do experimento. O que pode sugerir uma prefer?ncias das c?lulas ? presen?a do rhGH no biomaterial presente no meio de cultura, principalmente na matriz de PLGA. Os pol?meros testados, apresentaram ades?o e prolifera??o celular, por?m, os biomateriais ? base de PLGA, principalmente associados ao rhGH pareceram demonstrar maior prolifera??o celular quando avaliada a diferen?a do crescimento de 24 para 72horas. O GH modificou a superf?cie do pol?mero, aumentando a rugosidade e microporosidade. Aspecto visualizado principalmente quando incorporado ao PLGA. Mais estudos s?o necess?rios para verificar alternativas de novos biomateriais para libera??o lenta de f?rmacos.

Made available in DSpace on 2015-04-14T14:51:30Z (GMT). No. of bitstreams: 1 451789.pdf: 1080623 bytes, checksum: 53f3af2f0b9cc2bca520b792d55bcc75 (MD5) Previous issue date: 2013-03-27 / Somatotrophin or growth hormone (GH) is a hormone synthesized and secreted by the anterior pituitary gland in all animals and their secretion is controlled by two neuropeptides: the growth hormone releasing factor (GRF), which increases GH synthesis and secretion and somatostatin (SRIF), which inhibits GH secretion. bGH has galactopoietic effect, which is known since the thirties. The mechanism of action of bGH involves a series of orchestrated changes in the metabolism of body tissues so that more nutrients can be used for milk synthesis and these changes allow the animal to achieve an increased milk yield while remaining healthy. Long term studies (10 12 weeks) demonstrated increase in milk yield up to 40% with no adverse effects in the treated cows. E. coli is currently the largest platform for expression of heterologous proteins and high cell-density culture techniques for culturing E. coli are designed to increase productivity of proteins. This system is widely used for production of various recombinant proteins available in the market. In this work were performed bioreactor cultivations using fed batch techniques, testing different feeding strategies and different IPTG induction times. At the end of the experiments the biomass obtained was 32.4 g/L using a linear feeding strategy. Approximately 2.5 mg of bGH were obtained from 1 g of wet cell using a purification protocol with only one chromatographic column. / Somatotropina ou horm?nio de crescimento trata-se de um horm?nio natural secretado pela gl?ndula pituit?ria. Sua secre??o ? controlada por dois neuropept?deos secretados pelo hipot?lamo, o fator liberador de somatotropina, que aumenta a secre??o de horm?nio de crescimento e a somatostatina que inibe a sua secre??o. O horm?nio de crescimento bovino possui efeito galactopoi?tico, o que ? conhecido desde a d?cada de 30. Seu mecanismo de a??o envolve uma s?rie de mudan?as no metabolismo, direcionando mais nutrientes para a s?ntese de leite, sendo que o aumento na produ??o de leite ? alcan?ado sem prejudicar a sa?de do animal. Resultados de pesquisa demonstram que o horm?nio de crescimento bovino pode aumentar a produ??o de leite de 6% a 40% e a efici?ncia de utiliza??o de alimentos em bovinos. A produ??o de leite no Brasil aumentou cerca de 5 vezes desde 1975 at? 2011, e estima-se que existam 23 mil vacas leiteiras no pa?s, demonstrando o potencial mercado para o uso da somatrotopina bovina. E. coli ? atualmente a maior plataforma para express?o simples de prote?nas heter?logas, principalmente por ser um sistema bem caracterizado. Cultivos de alta densidade s?o desenvolvidos para aumentar a produtividade de prote?nas, sendo amplamente utilizados para produ??o de diversas prote?nas recombinantes dispon?veis no mercado. Neste trabalho foram realizados cultivos de E. coli em biorreator por meio de t?cnicas de batelada alimentada, testando diferentes estrat?gias de alimenta??o e tempos de indu??o com IPTG. Ao final dos experimentos a biomassa obtida foi de 32,4 g/L usando uma estrat?gia de alimenta??o linear. Aproximadamente 2,5 mg de bGH foram obtidas a partir de 1 g de c?lula ?mida usando um protocolo de purifica??o com apenas uma coluna cromatogr?fica.