Naslov
EFFECTS OF AUTOLOGOUS INTERLEUKIN 1 RECEPTOR ANTAGONIST PROTEINE ON ACHILLES TENDON HEALING IN RABBITS

Autori
Pećin, Marko

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
VII. MEĐUNARODNI KONGRES VETERINARSKA ZNANOST I STRUKA - Zbornik sažetaka / - Zagreb, 2017

Skup
VII. MEĐUNARODNI KONGRES VETERINARSKA ZNANOST I STRUKA - DAN DOKTORATA

Mjesto i datum
Zagreb, Hrvatska, 05.10.2017. - 07.10.2017

Vrsta sudjelovanja
Pozvano predavanje

Vrsta recenzije
Međunarodna recenzija

Ključne riječi
Achilles tendon ; healing ; interleukin-1 receptor antagonist protein ; interleukin-1 ; rabbit

Sažetak
The cytokine interleukin-1, more precisely its beta form (IL-1β), plays an important role in the initial stages of the inflammatory response in almost all tissues including tendons themselves. It is involved in the activation of inflammatory factors cascade and other molecules and also directly or indirectly increases its own local and systemic concentration by positive feedback. Recent studies are trying to work into this pro inflammatory cytokines by reducing or inhibiting completely its effects in inflammatory response. Precisely guided by the latest literature data demonstrating an important role of IL-1 in inflammation of tendons we decided to explore possibilities of preventing and improving treatment of inflammation of the Achilles tendon in rabbits by adding IL-1β (IL-1Ra) receptor antagonist protein from autologous conditioned serum. In this paper the assumption was that after longitudinal tenotomy of Achilles tendon inflammation will follow resulting in an increased concentration of IL-1β which will cause degradation of collagen fibers and extracellular matrix. Also it was expected that local administration of the therapeutic agent Orthokin (IL-1Ra, IRAP ®) in the first 48 hours after injury, will decrease the concentration of IL-1 in tendons. One of the hypothesis was that the Achilles tendon tissue of irap experimental group of rabbits, four weeks after the application of the antagonist histologically will be more similar to normal healthy tissue without pronounced inflammatory changes and without histological signs of chronic inflammation and degeneration in relation to the tissue of the control group. Considering the stated facts aim of this study was to determine the effect of autologous interleukin- 1 receptor antagonist protein (IL- 1Ra, IRAP ® Orthokin) on reducing iatrogenic created inflammation of the Achilles tendon in rabbits and its effect on the overgrowth of the same tendon 4 weeks after scarification and longitudinal tenotomies using histological and immunoenzymatic indicators. Also, we wanted to determine the amount of IL-1 with ELISA kit for rabbit IL-1 in tendon tissue and serum four weeks after the administration of antagonists. The goal was to measure residual inhibitory effect of IRAP® IL-1Ra on IL-1β and to determine the residual effect of the aforementioned doses of antagonists in the prevention of chronicity of the inflammatory process. The work was carried out on 26 Californian white rabbits, aged about 1-2 years and of both sexes. Rabbits were randomly divided into two equal groups (irap and control). The Achilles tendons of rabbits were used as a model. In both groups of rabbits tendons were prepared and dissected from the surrounding tissue. After tendon sheaths were opened tenotomies and scarification were performed using surgical scalpel no.15 along the entire length of tendon in the direction of the fibers. Longitudinal tenotomies resulted in the inflammation of the tendon. After closing of tendon sheaths in irap group of animals autologous conditioned serum rich with IL-1 receptor antagonist protein (IRAP ®, Orthokin) was injected into tendon in three points. In the control group buffered saline was administered with an equal amount, equal injection site and equal intervals as serum in the irap group (0.2 ml per injection site and 0, 24 and 48 hours after tenotomies). Four weeks after administration of the active components animals were sacrificed and the Achilles tendon were completely taken for morphological measurements, histological analysis and measurement of the amount of IL-1β by ELISA test kit for rabbit IL- 1β. Worked parameters included morphological and histological differences between experimental groups (irap and control) of animals differences were compared to normal physiologically tissue of the intact tendons. Appearance, layout, the number of collagen fibers, cellularity, the amount of glycosaminoglycans (GAGs) and elastic fibers were compared. Histological parameters of healing were assessed using specific staining and Bonar gradation scale from 0-3 with a statistically significant difference in irap group. One hundred µl of supernatant per one Achilles tendon was obtained after tissue underwent grinding, lysis and centrifugation and supernatant was used for ELISA. ELISA test was read at 450nm and concentration of IL-1 was determined using standard curve. More pronounced changes were observed in the control group such as tendon degeneration of collagen, improper orientation of the fibers, fiber thinning, hyper cellularity, scattered capillaries and an increased amount of GAG between the fibers. Four weeks postoperatively there was a distinct overall tendon-like appearance with thick collagen bundles and cells oriented along the tension fibers in tendons exposed to irap treatment. Tendons were thicker, had more unique and characteristic collagen fiber arrangement and more histologic maturity of the repair tissue than in control group. There were also differences in tendon healing of different parts of tendon tissue. We found increased amount the of IL-1β by 2.5 fold in the tissue and tendons of control group with negligible increase in the blood serum of the control group after 4 weeks. Difference in tissue samples histology and IL-1 concentration between the PBS and IL-1Ra group was statistically significant at week 4. There were also some statistically important differences between groups in platelet and white blood cell count. Increase in platelet number was noted in all control group rabbits. Neutropenia was noted in irap group. Inhibitory effect of IL-1Ra on the IL-1 prevented binding of IL-1 on to target cells surface of the tendons and consequently resulted in reduction of inflammation and degradation of collagen and extracellular matrix with histologically proven accelerated tendons healing. We concluded that IL-1Ra prevented morphological deterioration of the Achilles tendon in irap group. With minimal side effects such as local irritation and neutropenia with no apparent deteriorated health of experimental animals the prevention and treatment of inflammation of the Achilles tendon in rabbits using autologous serum rich with IL-1β receptor antagonist protein showed a good alternative for commonly used NSPU drugs and corticosteroids. Our study demonstrated that treatment of Achilles tendon iatrogenic inflammation with irap has the potential to improve tendon healing and could be considered as a treatment modality for Achilles tendon inflammation.

Izvorni jezik
Engleski

Znanstvena područja
Veterinarska medicina

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