Lesions of the distal deep digital flexor tendon (DDFT) are frequently diagnosed using magnetic resonance imaging (MRI) in horses with foot pain. The prognosis for horses with DDFT lesions to return to previous levels of performance is poor. Treatment options are limited; consisting of conservative therapy, desmotomy of the accessory ligament of the deep digital flexor tendon, injection of the digital sheath or navicular bursa, navicular bursoscopy or intralesional injection. Intralesional injection of biologic therapeutics shows promise in tendon healing, with increased number of experimental and clinical studies finding positive results. However, accurate injection of DDFT lesions within the hoof is difficult and requires general anesthesia. The Hallmarq open, low-field MRI unit was used to develop an MRI-guided technique to inject structures within the hoof. This procedure has been previously reported for injecting the collateral ligaments of the distal interphalangeal joint. Four clinical cases of deep digital flexor tendinopathy have been treated with MRI-guided injections using a similar technique. The aim of this study was to evaluate accuracy of a technique for injection of the deep digital flexor tendon within the hoof using MRI-guidance, which could be performed in standing patients. We hypothesized that injection of the DDFT within the hoof could be accurately guided using open low-field MRI to target either the lateral or medial lobe at a specific location. Ten cadaver limbs were positioned in an open, low-field MRI unit to mimic a standing horse. Each DDFT lobe was assigned to have a proximal (adjacent to the proximal aspect of the navicular bursa) or distal (adjacent to the navicular bone) injection. A titanium needle was inserted into each tendon lobe, guided by T1-weighted transverse images acquired simultaneously during injection. Oil-based colored dye was injected as a marker. Post-injection MRI and gross sections were assessed by three blinded investigators experienced in equine MRI. The success of injection as evaluated on gross section was 85% (70% proximal, 100% distal). The success of injection as evaluated by MRI was 65% (60% proximal, 70% distal). There was no significant difference between the success of injecting the medial versus lateral lobe. The major limitation of this study was the use of cadaver limbs with normal tendons. The authors concluded that injection of the DDFT within the hoof is possible using MRI guidance. Future work should be focused on using the techniqe in live horses with tendon lesions, and more clinical studies are needed to determine the most efficacious biologic therapeutic for tendon healing. / Master of Science

Orientador: Edson Rosa Pimentel / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-17T20:41:39Z (GMT). No. of bitstreams: 1 Vieira_CristianoPedrozo_M.pdf: 2180870 bytes, checksum: 84839495dd54341c9219c9aca477f0aa (MD5) Previous issue date: 2011 / Resumo: Os tendões podem ser acometidos por lesões, infecções e inflamações, seguidas ou não de ruptura, podendo ser decorrentes de atividades desportivas, como exercícios e alongamentos, ou de atividades diárias de muitos trabalhadores. Em situações patológicas a matriz extracelular (MEC) do tendão passa por um processo de reorganização de seus componentes, visando à regeneração e homeostasia do tecido. A inflamação pode ser desencadeada por diferentes fatores, entre os principais causadores desse processo estão injúrias mecânicas e químicas, agentes infecciosos, queimadura, radiação e supressão de oxigênio. Pouco é conhecido na literatura sobre as possíveis alterações que a inflamação instalada em tecidos próximos pode ocasionar em tendões. Desse modo, o presente estudo teve por objetivo analisar as alterações bioquímicas e morfológicas do tendão flexor digital profundo (TFDP) após indução da inflamação aguda em pata. Os períodos de análises foram 4 horas, período em que ocorre o pico da inflamação, 12 e 24 horas. Ratos Wistar (140-160g) foram separados em três grupos experimentais: os que receberam aplicação da carragenina (1%), os que receberam NaCl (0,9%), e os que não receberam nada, sendo utilizados como controle. O TFDP foi dividido conforme suas diferentes regiões (distal, intermediária, proximal). Para análises bioquímicas, os tendões foram processados e analisados de acordo com as seguintes técnicas: SDS-PAGE, para observação do perfil de proteínas, eletroforese em gel de agarose para análise de glicosaminoglicanos sulfatados; zimografia para detecção de metaloproteínase (MMP) 2 e 9; e dosagem de proteínas não colagênicas e hidroxiprolina. Para análises morfológicas, os cortes foram corados com hematoxilina-eosina, azul de toluidina e ponceau SS. De acordo com nossos resultados, no pico da inflamação aguda foi observada menor quantidade de proteínas e glicosaminoglicanos nas três regiões do TFDP dos animais tratados com carragenina. A concentração de hidroxiprolina foi maior nas duas regiões de tensão do tendão do grupo inflamado. A presença da MMP-9 foi detectada na região distal e foi evidenciado o epitendão mais espesso com células inflamatórias nas três regiões do TFDP no grupo com carragenina. Uma melhor organização dos feixes de colágeno foi observada nas duas regiões de tensão desse mesmo grupo. Após o período de pico da inflamação foi evidenciado a presença da isoforma latente e ativa da MMP-9 em 12 horas após a indução da inflamação. Houve maior quantidade de hidroxiprolina na região intermediária e proximal no grupo de 12 horas e, na região distal no grupo de 24 horas no grupo tratado com carragenina. A concentração de proteínas foi menor na região distal do grupo tratado com carragenina em 12 horas e maior em 24 horas nessa mesma região e grupo. A presença de um epitendão mais espesso com infiltrado de células foi observado nas regiões do TFDP dos animais com carragenina em 24 horas e, a organização dos feixes de colágeno foi menor na região proximal em 12 e 24 horas foram mostradas nos animais que receberam o veículo e a carragenina. Nenhuma diferença foi encontrada durante 4, 12 e 24 horas nos géis de SDS-PAGE. Nossos resultados mostram que embora o tendão não esteja inflamado, durante o pico do processo inflamatório agudo na pata de rato, alterações marcantes são evidenciadas. Contudo, podem ser ressaltados que o período posterior ao pico da inflamação também desencadeia alterações nos elementos estruturais e bioquímicos da MEC do TFDP / Abstract: The tendons are often affected by injuries, infections and inflammations, followed or not by rupture, which may occur during sport activities such as exercise and stretching, or during other daily activities. In pathological situations the extracellular matrix of tendons undergoes a reorganization process of their components, aimed at the improvement, regeneration and tissue homeostasis. Inflammation can be triggered by different factors, among the main causes of this process are mechanical and chemical injuries, infectious agents, burns, radiation and suppression of oxygen. Little is known in the literature on possible changes that the inflammation may trigger in near tissues where it is installed. This study aims to examine biochemically and morphologically the deep digital flexor tendon (DDFT) after induction of acute inflammation in rat paw (140-160g). The analysis was performed in the peak of inflammation (4 hours) and after that period (12 and 24 hours). Rats Wistar were divided into three groups: those who received application of (1%) carrageenan, those receiving 0.9% NaCl, and those who received nothing and were used as control. The DDFT was analyzed according to their regions (distal, intermediate and proximal). The DDFT was analyzed according to its different regions (distal, intermediate, proximal). For biochemical analysis, the tendons were processed and analyzed in accordance with the following techniques: SDS-PAGE, to observe the profile of proteins, agarose gel electrophoresis to analysis of sulfated glycosaminoglycans; zymography for detection of metalloproteinases 2 and 9, and dosage of non collagenous proteins and hydroxyproline. For morphological analysis, sections were stained with hematoxylin-eosin, toluidine blue and Ponceau SS. At the peak of acute inflammation was observed lower amount of proteins and glycosaminoglycans in the three regions of animals tendons with carrageenan. The hydroxyproline concentration was higher in the two tension regions of tendon of inflamed group. The presence of MMP-9 was detected in the distal region and was shown a thicker epitendon with inflammatory cells in the three regions of the DDFT in the group with carrageenan. Better organization of collagen bundles were observed within two regions of tension in the mentioned group. After the peak of inflammation was evidenced the presence of the latent and active isoform of the MMP-9 in 12 hours after induction of inflammation. A higher amount of hydroxyproline was detected in the intermediate and proximal region in the 12 hours and in the distal region in 24 hours in group treated with carrageenan. The protein concentration was lower in the distal region of the inflamed group at 12 hours and higher in 24 hours on the same region in the group treated with carrageenan. The presence of a thicker epitendon with cell infiltration was observed in the animals with DDFT of carrageenan animals as well as, a smaller organization of collagen bundles in the proximal region in 12 and 24 hours were shown in rats treated with vehicle and carrageenan. No difference was found for 4, 12 and 24 hours in SDS-PAGE gels. Our results show that although the tendon is not inflamed, during the peak of acute inflammation in rat paw, the most marked changes are evident. However, it can be emphasized that the period after the inflammation also triggers changes in the structural and biochemical components of the extracellular matrix of the deep digital flexor tendon / Mestrado / Biologia Celular / Mestre em Biologia Celular e Estrutural

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