Advancing Bone-Anchored Prostheses: Pros and Cons of Transtibial Osseointegration in Rabbits

Advancing Bone-Anchored Prostheses: Pros and Cons of Transtibial Osseointegration in Rabbits | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Advancing Bone-Anchored Prostheses: Pros and Cons of Transtibial Osseointegration in Rabbits Vladimir Ermishin, Soghoyan Gurgen, Dolgushkin Dmitry, Lebedev Mikhail, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-9539880/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Osseointegrated, percutaneous attachment can address limitations of socket prostheses but requires validated animal models to assess infection risk and long‑term stability. We developed and evaluated a transtibial osseointegration procedure in Soviet Chinchilla rabbits. Sixteen animals were enrolled in a two‑stage protocol. Stage one comprised tibial osteotomy, placement of a conical, self‑tapping, additively manufactured titanium intramedullary sleeve positioned flush with the cut surface, smoothing of bone edges, and coverage with a posterior muscle flap within a two‑flap skin–muscle approach. Stage two involved percutaneous pin insertion through a small incision with purse‑string stoma formation. Mean operative time was 40 ± 10 minutes with no intraoperative deaths and stable radiographic fixation. An initial pilot series (n = 5) revealed shortcomings – irregular osteotomy, soft‑tissue injury, and sleeve migration – that were mitigated by oscillating‑saw cuts under saline cooling, canal irrigation without aggressive reaming, and the revised flap design preserving posterior musculature. Complete stoma sealing remained challenging due to high skin mobility, and some postoperative complications, including fatalities, occurred. These findings provide a foundation for further research and for the evaluation of infection-control strategies, surgical techniques, and implant designs aimed at long-term outcomes. Orthopedics osseointegration bone-anchored prosthesis transtibial amputation percutaneous implant rabbit intramedullary titanium implant stoma formation Figures Figure 1 Figure 2 Introduction According to World Health Organization estimates about 40 million people worldwide live with limb amputations and this number continues to grow [1]. Conventional socket prostheses, which anchor to the skin often fail to meet patients’ demands for full functional recovery and work capacity due to numerous complications [2, 3]. An alternative is osseointegration – the approach introduced by Per-Ingvar Brånemark – which enables percutaneous bone-anchored prostheses [4, 5]. Initially applied in orthopedics and oncological surgery, this approach has evolved into percutaneous osseointegrated limb prosthetics, offering greater functional activity and quality of life than socket-based systems [6, 7]. Yet, problems exist with current osseointegrated prostheses, most importantly issues with infection risks and long-term durability, which necessitates comprehensive preclinical studies in animal models [8, 9]. In this study, we developed and tested a transtibial amputation model with percutaneous osseointegrated prosthetic implantation in soviet chinchilla rabbits. Materials and methods The study was carried out at the Research Institute of BioTech at Samara State Medical University (STMU). The study involved 16 Chinchilla rabbits – an average body weight of 5 ± 0.3 kg and age of 12–18 months old. The animal study protocol was reviewed and approved by the Bioethics Committee of STMU prior to study initiation (Protocol No. 297). All procedures complied with applicable legislation on the use of animals in research and relevant GLP requirements. The study also adhered to internationally accepted standards, including Directive 2010/63/EU, the Guide for the Care and Use of Laboratory Animals (8th ed., National Research Council), and the ARRIVE 2.0 reporting guidelines. Human Ethics and Consent to Participate declarations: not applicable. Animals were quarantined for 14 days, treated for parasites, and maintained on a standard diet with free water access. After surgery, they were housed individually and received a complete rabbit feed with hay. Feeding was carried out using a complete pellet diet designed for rabbits (non-pathogenic, maintenance diet). During the first three days after the surgery, to prevent gastrointestinal dysfunction, the animals were fed through an esophageal tube by the vivarium staff. If a rabbit refused hay or had no stool for more than 24 hours, it was given Bobotik 3 drops (produced by Polpharma, Poland) orally at a dose of 66.66 mg/mL. Implant Design To design the implants, hind limb radiographs of healthy rabbits were analyzed, and the tibial intramedullary canal width at the mid and distal thirds was measured via the digital ruler. Four sizes of conical intramedullary sleeves and a pin were developed. The sleeves had proximal grooves and dual threading: external self-cutting threads for insertion and internal threads for later pin attachment. Implants were produced from PTN-8 titanium powder (VT6, 10–45 µm) via Selective Laser Melting on a TruPrint 2000 (TRUMPF, Germany) 3D printer and sterilized by autoclaving at 180°C for 60 minutes (Fig. 1). Figure 1. Osseointegrated implant components. Left: conical intramedullary sleeve designed for tibial insertion. Right: percutaneous fixation pin for external prosthetic attachment. Preoperative Preparation Before surgery the hind limb was shaved. The animal was placed in the supine position on the operating table and the limbs were additionally secured. Surgical procedures were carried out in compliance with aseptic and antiseptic techniques, using intramuscular anesthesia. A mixture of Zoletil 100 (Virbac C.A., France) at 15 mg/kg body weight and Rometar (Bioveta, Czech Republic) at 6 mg/kg body weight was administered. Surgical Protocol The first stage began with an incision through the skin and subcutaneous tissue to create two full-thickness skin–fascial–muscle flaps (Fig. 2A). The anterior flap, based on the anterior tibial border, included fascia and anterior compartment muscles, while the larger posterior flap retained excess posterior compartment muscle. Hemostasis was achieved intraoperatively, and major vessels were ligated with nylon sutures. Figure 2. Surgical procedure for transtibial osseointegrated prosthetic implantation. A — Surgical approach and flap formation. B — Tibial osteotomy performed with an oscillating saw under continuous cooling. C — Insertion of the intramedullary sleeve using a flat screwdriver. D — Distal end of the sleeve positioned at the level of the bone cut. E — Resulting stump following the first stage of prosthetic surgery. F— Postoperative radiograph showing stable sleeve fixation with the distal end aligned to the tibial osteotomy level. G — Appearance of the stump after the second stage of surgery. An oscillating saw was used to perform an osteotomy 3 cm proximal to the flaps; the fibula was cut slightly more proximally than the tibia (Fig. 2B). The saw blade was cooled with saline to prevent thermal injury, and bone edges were smoothed with a rasp to reduce soft tissue trauma. The intramedullary canal was irrigated with sterile saline, and a self-tapping titanium implant was inserted flush (Fig. 2C) with the tibial cut surface (Fig. 2D). The posterior muscle flap was then positioned over the stump (Fig. 2E) and implant to form a muscular buffer, and antagonistic muscles were sutured with 3/0 Vicryl. In the second stage, a small skin incision exposed the implant end, which was cleaned; a pin was screwed into its sleeve, and the skin was closed with a purse-string suture to form a stoma (Fig. 2F). Postoperatively, radiographs were acquired in anteroposterior and lateral projections. Postoperative Care A veterinarian performed daily physical examinations and dressing changes. Examination included, evaluation of pain level via multidimensional Bristol Rabbit Pain Scale (BRPS) [9], and assessment of rectal body temperature, wound status and general condition. Wound status was estimated via edema of stump soft tissues, muscle hypotrophy, discharge, suture integrity, weight-bearing ability of the residual limb, range of motion in adjacent joints. General conditions included measurement of appetite, stool and sleep. Results In the 14 operated rabbits, no intraoperative deaths occurred; the mean duration of surgery was 40 ± 10 minutes. The finalized approach introduced a novel skin–muscle flap design that minimized postoperative trauma and reduced necrosis rates. Use of an oscillating saw with saline cooling prevented thermal bone injury and improved osteotomy quality, while a self-tapping sleeve provided stable fixation in the medullary canal, reducing implant migration. A preliminary study on five rabbits helped refine the protocol, including surgical access, instrumentation, and stump formation. Initial attempts lacked clear flap contouring and involved circumferential muscle dissection without preserving posterior muscle, leaving the bone edge exposed and causing wound perforation and infection. A Gigli saw produced debris, slippage, and irregular cuts, while unprocessed bone edges and an intact fibula caused soft tissue injury. Early canal reaming with a dental burr damaged cortical walls and caused sleeve migration. These complications were largely eliminated through protocol and sleeve design optimization. Conclusion Osseointegration represents one of the most promising frontiers in restorative and reconstructive medicine, offering the potential for seamless biomechanical and neurophysiological coupling between artificial and biological tissues. A major challenge remained achieving complete stoma sealing, hindered by high skin mobility and the lack of subcutaneous fat. Despite these improvements, some animals developed complications, including fatalities – consistent with reported mortality rates up to 18% and chronic complication rates of 59% [10]. Therefore, controlled animal studies remain essential to elucidate the biological mechanisms of bone–implant remodeling and to develop next-generation biomaterials and implant designs that ensure durable and functional integration. While rabbits have been recognized as a suitable model for osseointegration research, detailed surgical protocols for implant placement remain lacking. This study presents the first stage of a preclinical investigation in Soviet Chinchilla rabbits to establish and optimize a transtibial osseointegration procedure. Declarations Funding This research received no external funding. References Marino M, Pattni S, Greenberg M, et al. (2015) Access to prosthetic devices in developing countries: pathways and challenges. In: Proceedings of the global humanitarian technology conference, Seattle, WA, 8–11 October, pp.45–51. New York: IEEE. Sinegub AV, Kovalenko DА, Chupryaev VА, Nikolaenko AN, Borisov AP. (2025) Complications of Osseointegrated Prostheses and Comparison of Quality of Life in Patients with Different Prosthetic Systems: A Review. Traumatology and Orthopedics of Russia. 31(2):178-189. https://doi.org/10.17816/2311-2905-17663 Hoyt BW, Walsh SA, Forsberg JA. (2020) Osseointegrated prostheses for the rehabilitation of amputees (OPRA): results and clinical perspective. Expert Rev Med Devices. 17(1):17-25. https://doi.org/10.1080/17434440. 2020.1704623. Li Y, Brånemark R. (2017) Osseointegrated prostheses for rehabilitation following amputation: The pioneering Swedish model. Unfallchirurg. 120(4):285-292. https://doi.org/10.1007/s00113-017-0331-4 Hagberg K, Brånemark R. (2001) Consequences of non‐vascular trans‐femoral amputation: A survey of quality of life, prosthetic use and problems. Prosthetics and orthotics international. 25 (3): 186 – 194. https://doi.org/10.1080 /03093640108726601 Al Muderis M, Khemka A, Lord SJ, Van de Meent H, Frölke JP. (2016) Safety of Osseointegrated Implants for Transfemoral Amputees: A Two-Center Prospective Cohort Study. J Bone Joint Surg Am. 98(11):900-9. https://doi.org/ 10.2106/JBJS.15.00808 Leijendekkers RA, van Hinte G, Frölke JP, van de Meent H, Nijhuis-van der Sanden MW, Staal JB. (2017) Comparison of bone-anchored prostheses and socket prostheses for patients with a lower extremity amputation: a systematic review. Disabil Rehabil. 39(11):1045-1058. https://doi.org/10.1080/ 09638288.2016.1186752 Scarano A, Khater AGA, Gehrke SA, Inchingolo F, Tari SR. (2024) Animal Models for Investigating Osseointegration: An Overview of Implant Research over the Last Three Decades. J Funct Biomater. 15(4):83. https://doi.org/10.3390/jfb15040083 Benato L, Murrell J, Knowles TG, Rooney NJ. (2021) Development of the Bristol Rabbit Pain Scale (BRPS): A multidimensional composite pain scale specific to rabbits (Oryctolagus cuniculus). PLoS One. 16(6): e0252417. https://doi.org/ 10.1371/journal.pone.0252417 Northrup NC, Barron GH, Aldridge CF, Powers LV, Greenacre CB, Hutcheson JD, Morrisey JK. (2014) Outcome for client-owned domestic rabbits undergoing limb amputation: 34 cases (2000-2009). J Am Vet Med Assoc. 244(8):950-5. https://doi.org/10.2460/javma.244.8.950 Additional Declarations The authors declare no competing interests. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-9539880","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":630228672,"identity":"fa094dcf-5098-446a-9d78-e5bb8c44f7e3","order_by":0,"name":"Vladimir Ermishin","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA3UlEQVRIiWNgGAWjYBADHgYJxgaGDyRrYZxBmj0SDAzMPMQoNDjAncDwcc82GfnZzW2Pbf7YRctPO2PAXNmGTwvvBsYZz27zGNw52G6c25acu+F2jgHjWTxaJBt4NzDzHABqkUhsk85tYM7dIA3U0khIyx+gFvkZQC0Wf+pz588moIWfAaiFAaiF4QZQCwPb4dyG24S0MPNuONgDchhQi2Rv23GgX9IKDjacw62Fjb1344MfB27by89Ifybx40810GHJGx82lOHWwsDMwHAAQxBTZBSMglEwCkYBSQAAg61RUArNqzwAAAAASUVORK5CYII=","orcid":"https://orcid.org/0009-0003-0625-2233","institution":"Samara State Medical University","correspondingAuthor":true,"prefix":"","firstName":"Vladimir","middleName":"","lastName":"Ermishin","suffix":""},{"id":630228673,"identity":"d79fd72a-60b1-422f-9ee7-258fa0675692","order_by":1,"name":"Soghoyan Gurgen","email":"","orcid":"","institution":"Skoltech","correspondingAuthor":false,"prefix":"","firstName":"Soghoyan","middleName":"","lastName":"Gurgen","suffix":""},{"id":630228674,"identity":"fc66d9ff-624a-420b-ade3-2a26e5870849","order_by":2,"name":"Dolgushkin Dmitry","email":"","orcid":"","institution":"Samara State Medical University","correspondingAuthor":false,"prefix":"","firstName":"Dolgushkin","middleName":"","lastName":"Dmitry","suffix":""},{"id":630228676,"identity":"4f27cd74-6aee-410b-b346-d9838077612d","order_by":3,"name":"Lebedev Mikhail","email":"","orcid":"","institution":"Moscow State University","correspondingAuthor":false,"prefix":"","firstName":"Lebedev","middleName":"","lastName":"Mikhail","suffix":""},{"id":630228678,"identity":"b11f40ca-61c4-4a87-9bd8-e957800a133b","order_by":4,"name":"Minnebaev Ruslan","email":"","orcid":"","institution":"Skoltech","correspondingAuthor":false,"prefix":"","firstName":"Minnebaev","middleName":"","lastName":"Ruslan","suffix":""},{"id":630228679,"identity":"2fb9b132-3231-47db-87d6-cf49c70f6806","order_by":5,"name":"Nefedova Irina","email":"","orcid":"","institution":"Samara State Medical University","correspondingAuthor":false,"prefix":"","firstName":"Nefedova","middleName":"","lastName":"Irina","suffix":""},{"id":630228682,"identity":"0f2cc298-f93d-44c5-9661-a25c81435402","order_by":6,"name":"Lapteva Elena","email":"","orcid":"","institution":"Samara State Medical University","correspondingAuthor":false,"prefix":"","firstName":"Lapteva","middleName":"","lastName":"Elena","suffix":""},{"id":630228683,"identity":"c816e5c7-2e66-432a-8086-5302e54c3a1f","order_by":7,"name":"Sharafutdinova Anastasia","email":"","orcid":"","institution":"Samara State Medical University","correspondingAuthor":false,"prefix":"","firstName":"Sharafutdinova","middleName":"","lastName":"Anastasia","suffix":""},{"id":630228686,"identity":"d180f568-350e-41d6-9ccf-cc546541c675","order_by":8,"name":"Nikolaenko Andrey","email":"","orcid":"","institution":"Samara State Medical University","correspondingAuthor":false,"prefix":"","firstName":"Nikolaenko","middleName":"","lastName":"Andrey","suffix":""}],"badges":[],"createdAt":"2026-04-27 10:00:04","currentVersionCode":1,"declarations":{"humanSubjects":false,"vertebrateSubjects":true,"conflictsOfInterestStatement":false,"humanSubjectEthicalGuidelines":false,"humanSubjectConsent":false,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":true},"doi":"10.21203/rs.3.rs-9539880/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-9539880/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":108012812,"identity":"91aa560e-7f9a-40c1-9b84-fc64d965d696","added_by":"auto","created_at":"2026-04-28 13:16:27","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":172605,"visible":true,"origin":"","legend":"\u003cp\u003eOsseointegrated implant components.\u003c/p\u003e\n\u003cp\u003eLeft: conical intramedullary sleeve designed for tibial insertion. Right: percutaneous fixation pin for external prosthetic attachment.\u003c/p\u003e","description":"","filename":"Fig1.png","url":"https://assets-eu.researchsquare.com/files/rs-9539880/v1/e5137d5c221c4c556e105f47.png"},{"id":108012813,"identity":"691cbe2b-ce1d-4dfd-b141-e0b0a36229aa","added_by":"auto","created_at":"2026-04-28 13:16:27","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":18482733,"visible":true,"origin":"","legend":"\u003cp\u003eSurgical procedure for transtibial osseointegrated prosthetic implantation.\u003c/p\u003e\n\u003cp\u003eA — Surgical approach and flap formation.\u003c/p\u003e\n\u003cp\u003eB — Tibial osteotomy performed with an oscillating saw under continuous cooling.\u003c/p\u003e\n\u003cp\u003eC — Insertion of the intramedullary sleeve using a flat screwdriver.\u003c/p\u003e\n\u003cp\u003eD — Distal end of the sleeve positioned at the level of the bone cut.\u003c/p\u003e\n\u003cp\u003eE — Resulting stump following the first stage of prosthetic surgery.\u003c/p\u003e\n\u003cp\u003eF— Postoperative radiograph showing stable sleeve fixation with the distal end aligned to the tibial osteotomy level.\u003c/p\u003e\n\u003cp\u003eG — Appearance of the stump after the second stage of surgery.\u003c/p\u003e","description":"","filename":"Fig2.png","url":"https://assets-eu.researchsquare.com/files/rs-9539880/v1/f2a5c406fe08b091301b73d0.png"},{"id":108013092,"identity":"7d8d13da-42e5-4f16-aa16-fc3fc4e6d525","added_by":"auto","created_at":"2026-04-28 13:17:19","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":17734228,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-9539880/v1/35ec87bf-4c7f-41c6-a906-77a7c7ff0fae.pdf"}],"financialInterests":"The authors declare no competing interests.","formattedTitle":"\u003cp\u003e\u003cstrong\u003eAdvancing Bone-Anchored Prostheses: Pros and Cons of Transtibial Osseointegration in Rabbits\u003c/strong\u003e\u003c/p\u003e","fulltext":[{"header":"Introduction","content":"\u003cp\u003eAccording to World Health Organization estimates about 40\u0026nbsp;million people worldwide live with limb amputations and this number continues to grow [1]. Conventional socket prostheses, which anchor to the skin often fail to meet patients\u0026rsquo; demands for full functional recovery and work capacity due to numerous complications [2, 3]. An alternative is osseointegration \u0026ndash; the approach introduced by Per-Ingvar Br\u0026aring;nemark \u0026ndash; which enables percutaneous bone-anchored prostheses [4, 5]. Initially applied in orthopedics and oncological surgery, this approach has evolved into percutaneous osseointegrated limb prosthetics, offering greater functional activity and quality of life than socket-based systems [6, 7]. Yet, problems exist with current osseointegrated prostheses, most importantly issues with infection risks and long-term durability, which necessitates comprehensive preclinical studies in animal models [8, 9]. In this study, we developed and tested a transtibial amputation model with percutaneous osseointegrated prosthetic implantation in soviet chinchilla rabbits.\u003c/p\u003e"},{"header":"Materials and methods","content":"\u003cp\u003eThe study was carried out at the Research Institute of BioTech at Samara State Medical University (STMU). The study involved 16 Chinchilla rabbits \u0026ndash; an average body weight of 5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.3 kg and age of 12\u0026ndash;18 months old. The animal study protocol was reviewed and approved by the Bioethics Committee of STMU prior to study initiation (Protocol No. 297). All procedures complied with applicable legislation on the use of animals in research and relevant GLP requirements. The study also adhered to internationally accepted standards, including Directive 2010/63/EU, the Guide for the Care and Use of Laboratory Animals (8th ed., National Research Council), and the ARRIVE 2.0 reporting guidelines.\u003c/p\u003e \u003cp\u003eHuman Ethics and Consent to Participate declarations: not applicable.\u003c/p\u003e \u003cp\u003eAnimals were quarantined for 14 days, treated for parasites, and maintained on a standard diet with free water access. After surgery, they were housed individually and received a complete rabbit feed with hay. Feeding was carried out using a complete pellet diet designed for rabbits (non-pathogenic, maintenance diet).\u003c/p\u003e \u003cp\u003eDuring the first three days after the surgery, to prevent gastrointestinal dysfunction, the animals were fed through an esophageal tube by the vivarium staff. If a rabbit refused hay or had no stool for more than 24 hours, it was given Bobotik 3 drops (produced by Polpharma, Poland) orally at a dose of 66.66 mg/mL.\u003c/p\u003e \u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eImplant Design\u003c/h2\u003e \u003cp\u003eTo design the implants, hind limb radiographs of healthy rabbits were analyzed, and the tibial intramedullary canal width at the mid and distal thirds was measured via the digital ruler. Four sizes of conical intramedullary sleeves and a pin were developed. The sleeves had proximal grooves and dual threading: external self-cutting threads for insertion and internal threads for later pin attachment. Implants were produced from PTN-8 titanium powder (VT6, 10\u0026ndash;45 \u0026micro;m) via Selective Laser Melting on a TruPrint 2000 (TRUMPF, Germany) 3D printer and sterilized by autoclaving at 180\u0026deg;C for 60 minutes (Fig.\u0026nbsp;1).\u003c/p\u003e \u003cp\u003e \u003cb\u003eFigure 1.\u003c/b\u003e Osseointegrated implant components.\u003c/p\u003e \u003cp\u003eLeft: conical intramedullary sleeve designed for tibial insertion. Right: percutaneous fixation pin for external prosthetic attachment.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003ePreoperative Preparation\u003c/h3\u003e\n\u003cp\u003eBefore surgery the hind limb was shaved. The animal was placed in the supine position on the operating table and the limbs were additionally secured. Surgical procedures were carried out in compliance with aseptic and antiseptic techniques, using intramuscular anesthesia. A mixture of Zoletil 100 (Virbac C.A., France) at 15 mg/kg body weight and Rometar (Bioveta, Czech Republic) at 6 mg/kg body weight was administered.\u003c/p\u003e\n\u003ch3\u003eSurgical Protocol\u003c/h3\u003e\n\u003cp\u003eThe first stage began with an incision through the skin and subcutaneous tissue to create two full-thickness skin\u0026ndash;fascial\u0026ndash;muscle flaps (Fig.\u0026nbsp;2A). The anterior flap, based on the anterior tibial border, included fascia and anterior compartment muscles, while the larger posterior flap retained excess posterior compartment muscle. Hemostasis was achieved intraoperatively, and major vessels were ligated with nylon sutures.\u003c/p\u003e \u003cp\u003e \u003cb\u003eFigure 2.\u003c/b\u003e Surgical procedure for transtibial osseointegrated prosthetic implantation.\u003c/p\u003e \u003cp\u003eA \u0026mdash; Surgical approach and flap formation.\u003c/p\u003e \u003cp\u003eB \u0026mdash; Tibial osteotomy performed with an oscillating saw under continuous cooling.\u003c/p\u003e \u003cp\u003eC \u0026mdash; Insertion of the intramedullary sleeve using a flat screwdriver.\u003c/p\u003e \u003cp\u003eD \u0026mdash; Distal end of the sleeve positioned at the level of the bone cut.\u003c/p\u003e \u003cp\u003eE \u0026mdash; Resulting stump following the first stage of prosthetic surgery.\u003c/p\u003e \u003cp\u003eF\u0026mdash; Postoperative radiograph showing stable sleeve fixation with the distal end aligned to the tibial osteotomy level.\u003c/p\u003e \u003cp\u003eG \u0026mdash; Appearance of the stump after the second stage of surgery.\u003c/p\u003e \u003cp\u003eAn oscillating saw was used to perform an osteotomy 3 cm proximal to the flaps; the fibula was cut slightly more proximally than the tibia (Fig.\u0026nbsp;2B). The saw blade was cooled with saline to prevent thermal injury, and bone edges were smoothed with a rasp to reduce soft tissue trauma. The intramedullary canal was irrigated with sterile saline, and a self-tapping titanium implant was inserted flush (Fig.\u0026nbsp;2C) with the tibial cut surface (Fig.\u0026nbsp;2D). The posterior muscle flap was then positioned over the stump (Fig.\u0026nbsp;2E) and implant to form a muscular buffer, and antagonistic muscles were sutured with 3/0 Vicryl.\u003c/p\u003e \u003cp\u003eIn the second stage, a small skin incision exposed the implant end, which was cleaned; a pin was screwed into its sleeve, and the skin was closed with a purse-string suture to form a stoma (Fig.\u0026nbsp;2F). Postoperatively, radiographs were acquired in anteroposterior and lateral projections.\u003c/p\u003e\n\u003ch3\u003ePostoperative Care\u003c/h3\u003e\n\u003cp\u003eA veterinarian performed daily physical examinations and dressing changes. Examination included, evaluation of pain level via multidimensional Bristol Rabbit Pain Scale (BRPS) [9], and assessment of rectal body temperature, wound status and general condition. Wound status was estimated via edema of stump soft tissues, muscle hypotrophy, discharge, suture integrity, weight-bearing ability of the residual limb, range of motion in adjacent joints. General conditions included measurement of appetite, stool and sleep.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eIn the 14 operated rabbits, no intraoperative deaths occurred; the mean duration of surgery was 40\u0026thinsp;\u0026plusmn;\u0026thinsp;10 minutes. The finalized approach introduced a novel skin\u0026ndash;muscle flap design that minimized postoperative trauma and reduced necrosis rates. Use of an oscillating saw with saline cooling prevented thermal bone injury and improved osteotomy quality, while a self-tapping sleeve provided stable fixation in the medullary canal, reducing implant migration.\u003c/p\u003e \u003cp\u003eA preliminary study on five rabbits helped refine the protocol, including surgical access, instrumentation, and stump formation. Initial attempts lacked clear flap contouring and involved circumferential muscle dissection without preserving posterior muscle, leaving the bone edge exposed and causing wound perforation and infection. A Gigli saw produced debris, slippage, and irregular cuts, while unprocessed bone edges and an intact fibula caused soft tissue injury. Early canal reaming with a dental burr damaged cortical walls and caused sleeve migration. These complications were largely eliminated through protocol and sleeve design optimization.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eOsseointegration represents one of the most promising frontiers in restorative and reconstructive medicine, offering the potential for seamless biomechanical and neurophysiological coupling between artificial and biological tissues. A major challenge remained achieving complete stoma sealing, hindered by high skin mobility and the lack of subcutaneous fat. Despite these improvements, some animals developed complications, including fatalities \u0026ndash; consistent with reported mortality rates up to 18% and chronic complication rates of 59% [10]. Therefore, controlled animal studies remain essential to elucidate the biological mechanisms of bone\u0026ndash;implant remodeling and to develop next-generation biomaterials and implant designs that ensure durable and functional integration.\u003c/p\u003e \u003cp\u003eWhile rabbits have been recognized as a suitable model for osseointegration research, detailed surgical protocols for implant placement remain lacking. This study presents the first stage of a preclinical investigation in Soviet Chinchilla rabbits to establish and optimize a transtibial osseointegration procedure.\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eFunding\u003c/h2\u003e \u003cp\u003eThis research received no external funding.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eMarino M, Pattni S, Greenberg M, et al. (2015) Access to prosthetic devices in developing countries: pathways and challenges. In: Proceedings of the global humanitarian technology conference, Seattle, WA, 8\u0026ndash;11 October, pp.45\u0026ndash;51. New York: IEEE.\u003c/li\u003e\n\u003cli\u003eSinegub AV, Kovalenko DА, Chupryaev VА, Nikolaenko AN, Borisov AP. (2025) Complications of Osseointegrated Prostheses and Comparison of Quality of Life in Patients with Different Prosthetic Systems: A Review. Traumatology and Orthopedics of Russia. 31(2):178-189. https://doi.org/10.17816/2311-2905-17663\u003c/li\u003e\n\u003cli\u003eHoyt BW, Walsh SA, Forsberg JA. (2020) Osseointegrated prostheses for the rehabilitation of amputees (OPRA): results and clinical perspective. Expert Rev Med Devices. 17(1):17-25. https://doi.org/10.1080/17434440. 2020.1704623.\u003c/li\u003e\n\u003cli\u003eLi Y, Br\u0026aring;nemark R. (2017) Osseointegrated prostheses for rehabilitation following amputation: The pioneering Swedish model. Unfallchirurg. 120(4):285-292. https://doi.org/10.1007/s00113-017-0331-4\u003c/li\u003e\n\u003cli\u003eHagberg K, Br\u0026aring;nemark R. (2001) Consequences of non‐vascular trans‐femoral amputation: A survey of quality of life, prosthetic use and problems. Prosthetics and orthotics international. 25 (3): 186 \u0026ndash; 194. https://doi.org/10.1080 /03093640108726601\u003c/li\u003e\n\u003cli\u003eAl Muderis M, Khemka A, Lord SJ, Van de Meent H, Fr\u0026ouml;lke JP. (2016) Safety of Osseointegrated Implants for Transfemoral Amputees: A Two-Center Prospective Cohort Study. J Bone Joint Surg Am. 98(11):900-9. https://doi.org/ 10.2106/JBJS.15.00808\u003c/li\u003e\n\u003cli\u003eLeijendekkers RA, van Hinte G, Fr\u0026ouml;lke JP, van de Meent H, Nijhuis-van der Sanden MW, Staal JB. (2017) Comparison of bone-anchored prostheses and socket prostheses for patients with a lower extremity amputation: a systematic review. Disabil Rehabil. 39(11):1045-1058. https://doi.org/10.1080/ 09638288.2016.1186752\u003c/li\u003e\n\u003cli\u003eScarano A, Khater AGA, Gehrke SA, Inchingolo F, Tari SR. (2024) Animal Models for Investigating Osseointegration: An Overview of Implant Research over the Last Three Decades. J Funct Biomater. 15(4):83. https://doi.org/10.3390/jfb15040083\u003c/li\u003e\n\u003cli\u003eBenato L, Murrell J, Knowles TG, Rooney NJ. (2021) Development of the Bristol Rabbit Pain Scale (BRPS): A multidimensional composite pain scale specific to rabbits (Oryctolagus cuniculus). PLoS One. 16(6): e0252417. https://doi.org/ 10.1371/journal.pone.0252417\u003c/li\u003e\n\u003cli\u003eNorthrup NC, Barron GH, Aldridge CF, Powers LV, Greenacre CB, Hutcheson JD, Morrisey JK. (2014) Outcome for client-owned domestic rabbits undergoing limb amputation: 34 cases (2000-2009). J Am Vet Med Assoc. 244(8):950-5. https://doi.org/10.2460/javma.244.8.950\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"Samara State Medical University","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"osseointegration, bone-anchored prosthesis, transtibial amputation, percutaneous implant, rabbit, intramedullary titanium implant, stoma formation","lastPublishedDoi":"10.21203/rs.3.rs-9539880/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-9539880/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eOsseointegrated, percutaneous attachment can address limitations of socket prostheses but requires validated animal models to assess infection risk and long‑term stability. We developed and evaluated a transtibial osseointegration procedure in Soviet Chinchilla rabbits. Sixteen animals were enrolled in a two‑stage protocol. Stage one comprised tibial osteotomy, placement of a conical, self‑tapping, additively manufactured titanium intramedullary sleeve positioned flush with the cut surface, smoothing of bone edges, and coverage with a posterior muscle flap within a two‑flap skin\u0026ndash;muscle approach. Stage two involved percutaneous pin insertion through a small incision with purse‑string stoma formation. Mean operative time was 40\u0026thinsp;\u0026plusmn;\u0026thinsp;10 minutes with no intraoperative deaths and stable radiographic fixation. An initial pilot series (n\u0026thinsp;=\u0026thinsp;5) revealed shortcomings \u0026ndash; irregular osteotomy, soft‑tissue injury, and sleeve migration \u0026ndash; that were mitigated by oscillating‑saw cuts under saline cooling, canal irrigation without aggressive reaming, and the revised flap design preserving posterior musculature. Complete stoma sealing remained challenging due to high skin mobility, and some postoperative complications, including fatalities, occurred. These findings provide a foundation for further research and for the evaluation of infection-control strategies, surgical techniques, and implant designs aimed at long-term outcomes.\u003c/p\u003e","manuscriptTitle":"Advancing Bone-Anchored Prostheses: Pros and Cons of Transtibial Osseointegration in Rabbits","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-04-28 13:13:22","doi":"10.21203/rs.3.rs-9539880/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"58f36e52-c037-4fd2-8bc3-3c1de13bfed5","owner":[],"postedDate":"April 28th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":67077272,"name":"Orthopedics"}],"tags":[],"updatedAt":"2026-04-28T13:13:22+00:00","versionOfRecord":[],"versionCreatedAt":"2026-04-28 13:13:22","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-9539880","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-9539880","identity":"rs-9539880","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}