Magnesium-based bone implants: Immunohistochemical analysis of peri-implant osteogenesis by evaluation of osteopontin and osteocalcin expression A. Bondarenko, 1 N. Angrisani, 2 A. Meyer-Lindenberg, 3 J. M. Seitz, 4 H. Waizy, 5 J. Reifenrath 2 1 Department of Pathology, Dnipropetrovsk State Medical Academy, Ukraine, Dnipropetrovsk, Zhovtneva Ploshcha 14, 49005 2 Small animal clinic, University of Veterinary Medicine Hannover, B€ unteweg 9, 30559 Hannover, Germany 3 Clinic for Small Animal Surgery and Reproduction, Centre of Clinical Veterinary Medicine, Faculty of Veterinary Medicine, Ludwig-Maximilians-Universitaet Muenchen, Veterinaerstr. 13, 80539 Muenchen, Germany 4 Institute of Materials Science, Leibniz Universit€ at Hannover, An der Universit€ at 2, 30823 Garbsen, Germany 5 Department of Orthopaedics, Hannover Medical School, Anna-von-Borries Straße 3, 30625 Hannover, Germany Received 20 July 2012; revised 7 May 2013; accepted 31 May 2013 Published online 21 June 2013 in Wiley Online Library (wileyonlinelibrary.com). DOI: 10.1002/jbm.a.34828 Abstract: The functions of some bone proteins, as osteopon- tin (OPN) and osteocalcin (OC), have been discovered by the latest studies. This fact suggests the possibility of their immunodetection to characterize peri-implant osteogenesis and implant impact on it. Cylindrical pins of Mg alloys (MgCa0.8, LAE442, ZEK100, LANd442) and titanium alloy (TiAl6V4) were implanted into the tibial medullae of 46 rab- bits. Each group was divided regarding to implant duration (3 and 6 months). Bone samples adjacent to the implants were decalcified and treated with routine histological and immunohistochemical protocols using OC and OPN- antibodies. OC was detected in matrix of compact bone, but very rarely in osteoid and bone cells. OPN was detected intra- cellularly and in osteoid. After 3 months, the highest level of both markers was found in titanium group, followed by LAE442-group. In contrast to LAE442 and TiAl6V4, the other Mg alloys showed increasing levels of OC after 6 months. Lower levels of OP and OC compared to the control group are related to the continuous implant degradation and insta- bility of bone-implant interface in early post-surgical period. Reduced marker’s expression in LAE442 and TiAl6V4 groups after 6 months may indicate stabilization of bone-implant interface and completion of peri-implant neo-osteogenesis. Declining characters of OC and OPN expression over the implantation time, as well as their lowest levels in late post- surgical term, suggest a more appropriate biocompatibility of LAE442, which therefore seems to be the most preferable of the tested materials for the use in orthopaedic applications. V C 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 102A: 1449– 1457, 2014. Key Words: biodegradable bone implant, bone remodelling, in vivo, rabbit, immunohistochemistry, osteocalcin, osteopontin How to cite this article: Bondarenko A, Angrisani N, Meyer-Lindenberg A, Seitz JM, Waizy H, Reifenrath J. 2014. Magnesium- based bone implants: Immunohistochemical analysis of peri-implant osteogenesis by evaluation of osteopontin and osteocal- cin expression. J Biomed Mater Res Part A 2014:102A:1449–1457. BACKGROUND Resorbable magnesium-based implants can be assumed more preferable to the conventionally used non-resorbable metal implants due to the absence of removal surgery requirements and their satisfactory mechanical properties in vitro. 1,2 However, the biocompatibility is a compulsory assumption and depends substantially on the corrosion rate. 3 Another major requirement for bone implants is the stimulation of bone formation to advance fracture healing or fill up bone defects. Accordingly, the current studies on magnesium-based implants focused on both, reduction of the corrosion rate 4 and stimulation of bone formation. 5 For this purpose, the detailed understanding of the peri-implant osteogenesis and its alteration under the influence of implants with different chemical composition is of utmost importance. The biological mechanism of bone-implant interaction seems quite complex, and obviously involves a large amount of different regulatory pathways. 6 Because host responses to implants are significantly influenced by the protein signals encountered by the osteoprogeni- tor=osteoblast receptors, efforts of biomaterial research have focused on the biological influence of different implant materials onto the release of such ligands and their receptors in the bone tissue. 7,8 Nowadays there are many well-known biological substances which can up- or, contrary, down regulate bone growth as well as bone healing and remodelling. 9 The functions of such bone matrix proteins as osteopontin (providing osteoclast migration and adhesion; OPN) and osteocalcin (regulation of mineralization; OC) suggest the important role of their Correspondence to: N. Angrisani; e-mail: nina.angrisani@tiho-hannover.de V C 2013 WILEY PERIODICALS, INC. 1449