Research ArticleExperimental Studies
Open Access

Comparative Evaluation of Biocompatibility Data from the Subcutaneous and the Calvaria Implantation Model of a Novel Bone Substitute Block

KIM BURCKHARDT, OLE JUNG, MILENA RADENKOVIĆ STOŠIĆ, SANJA STOJANOVIĆ, STEVO NAJMAN, SVEN PANTERMEHL, REINHARD SCHNETTLER and MIKE BARBECK
In Vivo November 2025, 39 (6) 3116-3127; DOI: https://doi.org/10.21873/invivo.14113

Abstract

Background/Aim: This study compares the biocompatibility of a novel hybrid bone substitute material based on biphasic granules in a type I collagen scaffold using both subcutaneous and calvarial implantation models, in accordance with DIN EN ISO 10993-6. Given the distinct biological environments, materials may exhibit different behaviors in connective versus bone tissue. The aim was to evaluate irritation scores and cellular tissue responses to better understand the material’s performance in both settings.

Materials and Methods: This study evaluated the biocompatibility and host tissue response of test materials in male Wistar rats through subcutaneous and calvaria implantation models, following DIN EN ISO 10993-6, assessing cellular responses, material degradation, and bone regeneration at 10-, 30-, and 60-days post-implantation.

Results: Subcutaneous implants elicited a stronger inflammatory reaction with higher counts of polymorphonuclear cells, lymphocytes, multinucleated giant cells, and plasma cells at day 10, alongside consistently elevated irritancy scores. In contrast, calvaria implants showed increased neovascularization, reflecting bone-specific regenerative processes. Although capsule formation and cellular infiltration were similar between models, material degradation and phagocytosis were significantly greater subcutaneously at day 60.

Conclusion: These results highlight the critical impact of implantation site on immune activation, vascularization, and biomaterial resorption, underlining the importance of model selection in preclinical biomaterial assessment.

Keywords:

Introduction

DIN EN ISO 10993-6 is a key standard in the biocompatibility evaluation of medical devices, specifically addressing the local effects after implantation (1). This standard provides guidance for the macroscopic and microscopic evaluation of tissue responses to implanted materials, focusing on histopathological analysis to assess parameters such as inflammation, fibrosis, neovascularization, and bone remodeling. The scoring according to ISO 10993-6 involves the semi-quantitative assessment of cellular and tissue reactions surrounding the implant. Histological sections are examined and scored typically on a 0 to 4 scale for various cell types (e.g., polymorphonuclear cells, lymphocytes, macrophages, giant cells) and tissue changes (e.g., necrosis, fibrosis, neovascularization). Scores are then used to calculate a total irritation or tissue response score, which helps classify the material’s biocompatibility as non-irritant, slightly irritant, moderately irritant, or severely irritant (1-3).

In the course of the development and approval of new medical devices such as bone substitute materials (BSM), both the subcutaneous and the calvaria implantation model in rats are of special interest. For the regulatory approval of BSM–especially in the EU under medical device regulation (MDR) or in the US under the food and drug administration (FDA) guidance–both models are frequently used in a stepwise preclinical testing strategy. Thereby, the initial biocompatibility and safety is assessed via subcutaneous implantation, while the functional evaluation in bone tissue is tested using the calvarial defect model or other load-bearing defect models, depending on the intended application. Data from these models are part of the technical documentation submitted for regulatory review, demonstrating that the material is safe, biocompatible, and effective in supporting bone regeneration. Together, they form a robust foundation for preclinical evaluation and support the pathway toward regulatory approval.

When comparing material behavior in connective tissue versus bone tissue, several physiological and histological factors contribute to different outcomes. For example, vascularization and regeneration differ, as connective tissue is generally more vascularized and has higher regenerative capability compared to bone tissue (4, 5). As a result, materials implanted in soft tissues may show a faster resolution of inflammation and better integration, potentially leading to lower irritation scores. Also, the immune response sensitivity might significantly differ as the immune environment in connective tissue tends to respond more sensitively to foreign bodies such as bone substitute material (BSM), possibly leading to higher initial inflammatory scores, especially for materials prone to leaching or degradation (6, 7). Moreover, the tissue response in connective tissue often results in fibrotic encapsulation of the implant, while the material in the bone is directly bonding with the surrounding bone matrix. Materials that are favorable in one environment may not perform equally in the other.

Due to the distinct biological environments, materials may behave differently in connective versus bone tissue. Understanding these differences is essential for interpreting biocompatibility data, guiding material selection, and predicting clinical performance in various anatomical contexts. In this context, the present study compares the biocompatibility testing outcomes based on the DIN EN ISO 10993-6 scoring system using both implantation models investigating the inflammatory tissue response and biocompatibility of a novel hybrid bone block material combining synthetic biphasic BSM granules with a matrix of type I collagen. This material composition offers several key advantages in regenerative applications. The granules provide osteoconductivity and a controlled resorption profile and support long-term volume maintenance (7-9). Additionally, the collagen matrix is mimicking the natural extracellular matrix, promotes cell attachment, and may stimulate early vascular and cellular infiltration, while it also enhances handling properties (10-12). This combination might present a versatile and ready-to-use alternative to biologized bone blocks (e.g., autografts or xenografts), without the associated risks of donor site morbidity, immune response, or ethical concerns. Overall, this hybrid material might support effective bone regeneration while improving clinical handling, even in contrast to pure granular materials.

Materials and Methods

Biomaterial. In the present study a new hybrid bone substitute material block made of synthetic biphasic hydroxyapatite/β-tricalcium phosphate (HA/β-TCP) granules (BCP) embedded in a naturally crosslinked porcine collagen matrix (maxresorb® flexbone, biotrics bioimplants AG, Berlin, Germany) composed of collagen type I was analyzed.

In vivo study histology and analysis. The in vivo implantation procedures and subsequent histopathological analyses were performed in accordance with established protocols as described by Barbeck et al. (3, 13-15). This study was conducted in partnership with the Faculty of Medicine at the University of Niš, Serbia, with ethical approval granted by both the Local Ethical Committee of the Faculty and the Veterinary Directorate of the Ministry of Agriculture, Forestry, and Water Management of the Republic of Serbia (approval number: 323-07-01762/2019-05/9; approval date: 01/03/2019). Five experimental animals were assigned to each implantation model and time point (n=5) to ensure reliability and comparability of results, resulting in a total of 30 male Wistar rats examined at 10-, 30- and 60-days post-implantation. All animals survived surgical procedures without incident, and no abnormalities were observed.

Histological preparation. Explants were processed using an automated tissue processor (SLEE medical GmbH, Nieder-Olm, Germany) for dehydration and preparation for plastic embedding. Stepwise infiltration at 4°C with Technovit 9100 medium was followed by polymerization according to the manufacturer’s instructions. Explants were positioned in rolled rim bottles (VWR, Darmstadt, Germany) with the cutting surface facing downward, filled with the polymerization mixture, sealed airtight, and stored at −20°C until hardened. Tissue blocks were trimmed (EcoMet 30, Buehler, Esslingen, Germany), sectioned at 4-6 μm (CUT4060E, microTec GmbH, Walldorf, Germany), and subjected to hematoxylin and eosin (HE) staining.

Histopathological analyses. To evaluate tissue compatibility and host response, histological examination focused on material integration within the implantation sites, in accordance with DIN EN ISO 10993-6 (1, 16). Multiple histopathological criteria were assessed to ensure the safety of the test materials, including the presence of granulocytes, lymphocytes, plasma cells, macrophages, and multinucleated giant cells, as well as tissue characteristics such as fibrosis, neovascularization, and adipose tissue infiltration.

Based on the frequency and distribution of these cellular and tissue responses, the slides were evaluated and classified using the irritancy/reactivity grading system outlined in Annex E of ISO 10993-6. The irritancy score for each test and control group was determined by calculating the average score of all corresponding implantation sites.

Furthermore, the parameters for resorbable materials, i.e., capsule formation, infiltration/inflammation, cell growth within the implant, degradation and phagocytosis, were evaluated in accordance with DIN EN ISO 10993-6 and the scoring system based on a publication by De Jong et al. (17). Finally, bone regeneration in the calvaria model was scored based on a scoring system introduced by Musson et al. (18).

Statistical analysis. The raw scoring data were analyzed statistically using the Mann–Whitney U test to compare two independent groups, employing GraphPad Prism version 10.2 (GraphPad Software Inc., La Jolla, CA, USA). Differences were considered statistically significant at p≤0.05 (*p≤0.05), highly significant at p≤0.01 (**p≤0.01), and very highly significant at p≤0.001 (***p≤0.001). Data are expressed as mean±standard deviation.

Results

Scanning electron microscopy (SEM). The SEM analysis revealed that both the biphasic BSM granules and the surrounding type I collagen scaffold were clearly visualized (Figure 1). The analysis indicated that the BSM granules are irregularly distributed within the collagen scaffold, exhibiting numerous pores with varying diameters and structural irregularities.

Figure 1.
Figure 1.

Bone substitute scaffold composed of synthetic biphasic bone substitute material (BS) granules integrated within a surrounding collagen scaffold (CS). Scanning electron microscopy (SEM) images illustrate the morphology and surface structure of the scaffold: left, overview at 100× magnification; right, higher resolution at 200× magnification, highlighting the porous architecture and close interaction between BS granules and CS matrix.

In vivo analysis. Histological analysis indicated that the biphasic bone substitute material (BSM) elicited granulation tissue formation at all study time points and in both implantation models (Figure 2). Macrophages and multinucleated giant cells were the primary cell types involved in the tissue response, with comparable numbers observed across tissues and time points (Figure 2). Although macrophage counts remained similar between implantation models, the number of giant cells was consistently higher in the subcutaneous connective tissue, reaching the greatest disparity on day 10 post-implantation. Additionally, polymorphonuclear cell numbers peaked at day 10 but were comparable between groups at 30 and 60 days. Lymphocyte counts were slightly elevated in the subcutaneous implantation bed compared to the calvaria implantation area throughout the study. Notably, moderate plasma cell infiltration was observed solely in the subcutaneous implantation beds of the biphasic bone substitute, with a decreasing trend over time. Both implantation models exhibited low levels of necrosis and vascularization throughout the study, though vascularization was marginally increased in the calvaria beds at day 10. Signs of fibrosis were minimal at 10 and 30 days in both study groups, while increased fibrosis was observed in the calvaria implantation beds at day 60 post-implantation. The collagen part of the biphasic BSM was detectable until 30 days post-implantation, while the main reactivity was induced by the synthetic HA/β-TCP granules (Figure 2).

Figure 2.
Figure 2.

Representative histological images of biphasic bone substitute (BS) scaffolds implanted in subcutaneous tissue (left column) and calvarial defects (right column) at 10 days (top row), 30 days (middle row), and 60 days (bottom row) post-implantation. Hematoxylin and eosin (H&E) staining at 400× magnification illustrates the host tissue response and cellular interactions with the biomaterial. BS: Bone substitute granules; CT: connective tissue. Black arrows indicate macrophages, blue arrows indicate lymphocytes, and yellow arrows indicate granulocytes, demonstrating immune cell infiltration. Red arrows highlight blood vessels, indicating neovascularization within the implantation beds. Black arrowheads mark multinucleated giant cells, while white arrowheads denote multinucleated giant cells containing phagocytosis vesicles. Scale bars=10 μm.

Within the calvaria implantation model, the process of bone regeneration displayed a dynamic and time-dependent pattern, characterized by a gradual and continuous increase in newly formed bone matrix. Over the course of the study, the amount and density of the matrix became progressively more pronounced, reflecting a steady maturation and integration of the regenerating tissue into the defect area (Figure 3). At day 10 post-implantation, new bone formation was observed primarily at the edges of the defect areas, with minimal osteoconductive guidance provided by the bone substitute granules (Figure 3A). By day 30 post-implantation, increased integration of the material granules into the bone regeneration process was detected, especially in the marginal regions (Figure 3B), while by day 60 post-implantation, the granules were nearly completely surrounded by new bone matrix (Figure 3C).

Figure 3.
Figure 3.

Representative histological images of calvarial implantation beds containing biphasic bone substitute (BS) scaffolds at 10 (A), 30 (B), and 60 (C) days post-implantation. Hematoxylin and eosin (H&E) staining at 400× magnification, scale bars = 10 μm. BS: Bone substitute granules; NB: newly formed bone matrix; CT: connective tissue; green arrows indicate osteoblasts. At 10 days (A), early bone formation is observed with sparse NB and initial osteoblast activity along the scaffold surfaces. By 30 days (B), the amount of NB has increased, showing a more organized structure and closer integration with the surrounding CT, accompanied by active osteoblasts depositing matrix. At 60 days (C), the NB appears more mature and densely packed, with osteoblasts still present along the bone surfaces, indicating ongoing remodeling and progressive incorporation of the scaffold into the host tissue.

The histopathological scoring resulted in the irritancy scores that revealed that the material after subcutaneous implantation had an average treatment irritancy score of 19.7, and the material implanted in the calvaria defects had an average treatment irritancy score of 15.0 at 10 days post-implantation. Thus, the difference between the irritancy scores in both implantation models was 4.7. At 30 days post-implantation the subcutaneously implanted material had an average treatment irritancy score of 19.9, and the material after calvaria implantation had an average treatment irritancy score of 15.4. Thus, the difference between the irritancy scores in both implantation models was 4.5. At 60 days post-implantation, the subcutaneous implants exhibited an average treatment irritancy score of 18.9, while the calvarial implants had an average score of 13.5. Therefore, the difference in irritancy scores between the two implantation models was 5.4.

Furthermore, a statistical analysis of the scoring data was conducted (Figure 4). The analysis revealed that the material induced statistically significant higher scoring data after subcutaneous implantation (p<0.05 and p<0.01) only at day 10 post-implantation in view of polymorphonuclear cells, lymphocytes and giant cells (Figure 4A, B, and E). Furthermore, significantly higher neovascularization scores (p<0.05) were detected in the calvaria implantation model at this early time point (Figure 4G). Regarding all other scoring parameters and the later study points, no further significant differences were found (Figure 4).

Figure 4.
Figure 4.

Quantitative results of the histological scoring analysis at 10-, 30-, and 60-days post-implantation, evaluated according to the DIN EN ISO 10993-6 standard. (A) Polymorphonuclear cells, (B) lymphocytes, (C) plasma cells, (D) macrophages, (E) giant cells, (F) necrosis, (G) neovascularization and (H) fibrosis. Scores reflect the tissue response to the biphasic bone substitute scaffolds, including inflammation, connective tissue formation, and integration with the host tissue. Significant differences (Mann–Whitney U-test) between the implantation tissues are indicated (*p<0.05, **p<0.01).

Additionally, the scoring of the parameters for resorbable materials, showed that no differences in capsule formation, infiltration/inflammation and cell growth within the implant were found throughout the study between both implantation models, although a slight increase was found in the calvaria implantation model at day 60 post implantation (Figure 5A-C). Regarding degradation and phagocytosis, only significant differences with higher values in the subcutaneous implantation model were found at day 60 post-implantation (*p<0.05 and ***p<0.001, respectively; Figure 5D and E). Interestingly, the scoring of bone regeneration showed that no differences over the study course were found throughout the study (Figure 6).

Figure 5.
Figure 5.

The scoring analysis workflow of bioresorbable implants was conducted at 10-, 30-, and 60-days post-implantation to evaluate their in vivo performance over time. (A) Capsule formation, (B) Infiltration/ inflammation, (C) Cell growth within the implant, (D) degradation and (E) phagocytosis. This analysis enabled a comprehensive assessment of tissue response, implant integration, and degradation kinetics at each time point. Statistically significant differences (Mann–Whitney U test) between experimental groups are indicated, with *p<0.05 representing significance and ***p<0.001 denoting highly significant differences.

Figure 6.
Figure 6.

Bone regeneration scores in the calvaria model at 10-, 30-, and 60-days post-implantation, illustrating temporal changes in new bone formation.

Discussion

Both the subcutaneous and the calvaria implantation model are standard models for the analysis of the biocompatibility of medical devices intended for bone repair. Due to the distinct biological environments, materials may behave differently in connective versus bone tissue. Understanding these differences is essential for interpreting biocompatibility data, guiding material selection, and predicting clinical performance in various anatomical contexts. The aim of the present study was to comparatively evaluate irritation scores and cellular tissue responses within the framework of biocompatibility testing according to DIN EN ISO 10993-6, using both implantation models. For this purpose, a novel hybrid bone substitute material composed of biphasic BSM granules embedded in a collagen type 1 scaffold was utilized.

The present study demonstrates clear differences in tissue response between the subcutaneous and calvaria implantation models, particularly at day 10 post-implantation. At this time point, significantly higher levels of polymorphonuclear cells, lymphocytes, and multinucleated giant cells were observed in the subcutaneous implantation group. This suggests a more pronounced inflammatory reaction in the subcutaneous tissue environment. In contrast, neovascularization was significantly more prominent in the calvaria implantation model at the same time point, indicating enhanced vascular response in the bone-associated site.

The elevated number of immune cells in the subcutaneous group may reflect a stronger foreign body reaction, which is often seen in soft tissue environments. Subcutaneous tissue is characterized by a highly vascularized and loose connective matrix that allows for more rapid recruitment of inflammatory cells following implantation (19). Polymorphonuclear cells are among the first responders in the acute inflammatory phase and their prolonged presence, alongside lymphocytes and multinucleated giant cells, may indicate the progression to chronic inflammation and a more active foreign body reaction. The formation of multinucleated giant cells is, in particular, a hallmark of the body’s attempt to degrade or isolate foreign materials and is often associated with persistent irritation or immune activation (19).

On the other hand, the calvaria model showed significantly greater neovascularization at day 10, suggesting that bone tissue provides a more conducive environment for angiogenic processes during the early healing phases. Bone is inherently more angiogenically active due to its need to maintain a constant blood supply for remodeling and regeneration. In the context of biomaterial implantation, the calvaria site can support rapid vascular ingrowth, potentially facilitated by mechanical stimulation and the presence of osteoprogenitor cells (20). Enhanced vascularization is critical for the successful integration and long-term functionality of biomaterials, especially those intended for bone regeneration, as it ensures adequate nutrient delivery, waste removal, and recruitment of progenitor cells (21).

A particularly interesting and novel finding is the exclusive presence of plasma cells in the subcutaneous implantation model. Plasma cells are terminally differentiated B lymphocytes responsible for antibody production, and their appearance is typically associated with a more advanced or chronic stage of the immune response. As no plasma cells were found in the calvaria model, it can be concluded that their presence in the subcutaneous implantation model must be related to stimuli other than the biomaterial. The detection of plasma cells exclusively in the subcutaneous group on the one hand suggests a more robust adaptive immune activation in this model. This observation supports the notion that subcutaneous implantation may lead not only to a heightened acute inflammatory response but also to a stronger engagement of the humoral immune system. The presence of plasma cells, in combination with increased levels of lymphocytes and multinucleated giant cells, may indicate the development of a foreign body granulomatous reaction or chronic inflammation in response to the implanted material. Chronic inflammatory responses involving plasma cells are often associated with persistent antigenic stimulation, as seen in biomaterial-related immune reactions or delayed clearance of degradation products (19). This may reflect the body’s attempt to mount a more sophisticated immune defense in the absence of osseous integration cues, which are more prominent in bone environments. Conversely, the primary distinction between the two models lies in the mechanical impact exerted on implanted biomaterials. The calvaria model provides a mechanically neutral environment for testing, whereas the calvarial model subjects the implant to low-level mechanical stimuli, partly due to the movement of experimental animals. While plasma cells are not classically considered mechanosensitive, growing evidence suggests their behavior is influenced by mechanical cues in their microenvironment. For example, changes in extracellular matrix (ECM) stiffness, and vascular dynamics can affect stromal cells, which in turn regulate plasma cell survival, localization, and antibody production via factors like CXCL12 and IL-6 (22, 23). In pathological settings, plasma cells show a preference for stiffer ECM regions, likely mediated by integrin and FAK signaling (24). In inflammatory or fibrotic tissues, increased stiffness may indirectly impact plasma cells by altering stromal support (25). In vitro models also suggest that tuning substrate mechanics can enhance plasma cell viability and function (26). Though direct mechanosensing by plasma cells remains unclear, mechanical forces clearly influence their niche and function through indirect mechanisms, which could also be a reason for their occurrence in the subcutaneous implantation beds in the present study.

In addition to the cellular and vascular differences observed, the scoring of specific tissue response parameters to resorbable materials further illustrates the nuanced interaction between implantation site and biomaterial behavior. Interestingly, no significant differences in capsule formation, overall cellular infiltration/inflammation, or cell ingrowth within the implant were detected between the subcutaneous and calvaria models throughout the observation period. These findings suggest that, despite distinct early immune profiles, the fundamental integration processes for resorbable materials follow similar trajectories in both tissue types. However, at the latest 60-day post-implantation time-point, a slight increase in these parameters was noted in the calvaria model, which may reflect prolonged or ongoing remodeling activity typical for bone environments (27-29).

Notably, significant differences were observed for the degradation and phagocytosis parameters at day 60, with markedly higher scores in the subcutaneous group. These results suggest a more advanced or accelerated breakdown of the biomaterial in the soft tissue environment, likely facilitated by the higher presence of phagocytic cells such as multinucleated giant cells and macrophages earlier in the response (19, 30). The enhanced degradation may also correlate with the earlier appearance of plasma cells and the stronger inflammatory stimulus in subcutaneous tissue. In contrast, the calvaria model, despite its higher neovascularization, appears to favor a more controlled degradation pathway, possibly linked to its more stable and mineralized environment (31). These differences emphasize the impact of implantation site not only on immune responses but also on material resorption dynamics, which is critical for the design of degradable biomaterials (29, 32, 33).

These findings underscore the importance of carefully selecting the implantation model when evaluating new biomaterials. While subcutaneous implantation provides a useful and accessible model for evaluating immunogenicity, tissue compatibility, and early inflammatory responses, it does not fully replicate the complex environment of bone tissue. In contrast, calvaria or other orthotopic models more closely mimic clinical bone repair scenarios, particularly in terms of vascular response and osteoconductive behavior.

The divergence in cellular response between these two models at day 10 suggests that the temporal dynamics of healing and immune modulation are strongly dependent on the implantation site. Therefore, interpretation of biocompatibility and regenerative potential must consider the specific microenvironment in which the material is tested. Future studies may benefit from combining both models in a complementary fashion – using the subcutaneous model to evaluate immunological safety and the calvaria model to assess functional integration and angiogenesis.

A further key finding was the analysis of treatment irritancy scores across time points. At all evaluated stages (days 10, 30, and 60), subcutaneous implants consistently exhibited higher irritancy scores compared to calvaria implants. At day 10, the difference between models was 4.7 (subcutaneous: 19.7; calvaria: 15.0). This trend continued at day 30 (difference of 4.5) and became even more pronounced by day 60 (difference of 5.4), indicating a sustained higher level of irritation in subcutaneous tissue. These data reinforce the interpretation that the subcutaneous environment provokes a stronger and longer-lasting host reaction, potentially due to reduced mechanical stability, persistent inflammatory cell infiltration, and a less regenerative microenvironment (19, 34).

Altogether, these findings underscore the importance of implantation site selection in preclinical biomaterial testing. The subcutaneous model is valuable for evaluating immunogenicity, phagocytosis, and irritancy potential, but may overestimate clinical inflammation levels. In contrast, the calvaria model provides a more relevant context for bone-related applications, particularly regarding angiogenesis and osseointegration. Combining both models may offer a holistic understanding of biomaterial behavior under diverse physiological conditions, aiding in the design of safer and more effective tissue-engineered constructs. Moreover, according to data from both implantation models, the hybrid biomaterial demonstrates biocompatibility, as the observed tissue responses are consistent in type and intensity with those reported for other bone substitute materials.

Conclusion

The comparative analysis of subcutaneous and calvarial implantation models reveals that the implantation site significantly influences the host tissue response to resorbable biomaterials. Subcutaneous implants provoke a stronger inflammatory and immune reaction, evidenced by higher polymorphonuclear cells, lymphocytes, plasma cells, and irritancy scores throughout the study period. In contrast, calvaria implants demonstrate enhanced neovascularization and more controlled degradation, reflecting the bone’s regenerative environment. These findings highlight the importance of selecting appropriate implantation models to accurately evaluate biomaterial biocompatibility and performance for specific clinical applications.

Acknowledgements

The authors gratefully acknowledge the funding by the German Research Foundation (Deutsche Forschungsgemeinschaft, DFG) for the research unit 5250 “Mechanism-based characterization and modeling of permanent and bioresorbable implants with tailored functionality based on innovative in-vivo, in-vitro and in-silico methods” (project no. 449916462). The Authors further thank the Chair of Materials Test Engineering (WPT), TU Dortmund University, Dortmund, Germany, for providing the SEM micrographs (Figure 1) within the framework of an excellent scientific collaboration.

Footnotes

  • Authors’ Contributions

    Conceptualization: O.J. and M.B.; methodology: K.B., O.J., M.R., S.S., S.S., S.P., R.S., and M.B.; in vivo analyses: K.B, O.J., S.p. and M.B.; software: M.B.; validation, K.B. and M.B.; formal analysis: K.B., O.J., M.R., S.S., S.S., S.P., R.S., and M.B.; investigation: K.B., O.J., M.R., S.S., S.S., S.P., R.S., and M.B.; resources: O.J., S.S., S.S. and M.B.; data curation: K.B., O.J., M.R., S.S., S.S., S.P., R.S., and M.B.; writing - original draft preparation: K.B., O.J., M.R., S.S., S.S., S.P., R.S., and M.B.; writing - review and editing: K.B. and M.B.; visualization: K.B., O.J., M.R., S.S., S.S., S.P., R.S., and M.B.; supervision: M.B.; project administration: M.B.; funding acquisition: O.J., S.S., S.N. and M.B. All Authors have read and agreed to the published version of the manuscript.

  • Conflicts of Interest

    All the Authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

  • Artificial Intelligence (AI) Disclosure

    No artificial intelligence (AI) tools, including large language models or machine learning software, were used in the preparation, analysis, or presentation of this manuscript.

  • Received August 5, 2025.
  • Revision received September 9, 2025.
  • Accepted September 15, 2025.

This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY-NC-ND) 4.0 international license (https://creativecommons.org/licenses/by-nc-nd/4.0).

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Comparative Evaluation of Biocompatibility Data from the Subcutaneous and the Calvaria Implantation Model of a Novel Bone Substitute Block
KIM BURCKHARDT, OLE JUNG, MILENA RADENKOVIĆ STOŠIĆ, SANJA STOJANOVIĆ, STEVO NAJMAN, SVEN PANTERMEHL, REINHARD SCHNETTLER, MIKE BARBECK
In Vivo Nov 2025, 39 (6) 3116-3127; DOI: 10.21873/invivo.14113
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