Multinucleated giant cells within the in vivo implantation bed of a collagen-based biomaterial determine its degradation pattern

Abstract

Objectives: The aim of the present study was to characterize the cellular reaction to a xenogeneic resorbable collagen membrane of porcine origin using a subcutaneous implantation model in Wistar rats over 30 days.

Materials and methods: Ex vivo, liquid platelet-rich fibrin (PRF), a leukocyte and platelet-rich cell suspension, was used to evaluate the blood cell membrane interaction. The material was implanted subcutaneously in rats. Sham-operated rats without biomaterial displayed physiological wound healing (control group). Histological, immunohistological, and histomorphometric analyses were focused on the inflammatory pattern, vascularization rate, and degradation pattern.

Results: The membrane induced a large number of mononuclear cells over the observation period, including lymphocytes, macrophages, and fibroblasts. After 15 days, multinucleated giant cells (MNGCs) were observed on the biomaterial surface. Their number increased significantly, and they proceeded to the center of the biomaterial on day 30. These cells highly expressed CD-68, calcitonin receptor, and MMP-9, but not TRAP or integrin-ß3. Thus, the membrane lost its integrity and underwent disintegration as a consequence of the induction of MNGCs. The significant increase in MNGC number correlated with a high rate of vascularization, which was significantly higher than the control group. Physiological wound healing in the control group did not induce any MNGCs at any time point. Ex vivo blood cells from liquid-PRF did not penetrate the membrane.

Conclusion: The present study suggests a potential role for MNGCs in biomaterial degradation and questions whether it is beneficial to accept them in clinically approved biomaterials or focus on biomaterials that induce only mononuclear cells. Thus, further studies are necessary to identify the function of biomaterial-induced MNGCs.

Clinical relevance: Understanding the cellular reaction to biomaterials is essential to assess their suitability for specific clinical indications and outline the potential benefit of specific group of biomaterials in the respective clinical indications.

Keywords: Disintegration; Guided bone regeneration (GBR); Guided tissue regeneration (GTR); Inflammatory pattern; Integration.

Fig. 1

Ex vivo interaction between membrane Creos™ Xenoprotect (CXP), blood-derived cells, and fibrin from liquid-PRF. a A control cross section of the native membrane ex vivo, H and E staining; × 100 magnification. b CXP after liquid-PRF application. A fibrin clot is formed on the membrane surface (*); H and E staining; × 100 magnification. c High magnification of fibrin clot formation on the membrane surface (*) including blood cells (arrows). The membrane was not invaded by cells

Fig. 2

The cellular reaction to Creos™ Xenoprotect (CXP). a and b Day 3. c and d Day 15. e and f day 30. Left column azan staining; × 100 magnification. Right column Masson-Goldner staining; × 200 magnification. Arrows, peri-implantation tissue/cells

Fig. 3

Immunhistological staining of the different expression markers in multinucleated giant cells (MNGCs). Positive cells are stained in red/brown. a Anti-CD-68. b Anti-calcitonin receptor. c Anti-MMP-9. d Anti-integrin-β3. e TRAP staining. f Statistical analysis of the histomorphometrically measured MNGC number per square millimeter. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001

Fig. 4

Biomaterial-induced multinucleated giant cells (MNGCs). aTRAP-negative MNGCs (arrowhead) on Creos™ Xenoprotect (CXP) surface on day 15. b TRAP-positive cells (arrowheads) invaded CXP on day 30. TRAP staining; × 200 magnification. c Statistical analysis of the histomorphometrically measured MNGC number per square millimeter. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001

Fig. 5

Vessel formation within Creos™ Xenoprotect (CXP). Statistical analysis of the histomorphometrically measured vessel number per square millimeter. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. (a) Vessel density, (b) vascularization rate as a percent, (c–e) immunohistological staining of α-SMA marked vessels (arrows) (c) day 3, (d) day 15, and (e) and (e1) day 30. All images were captured at × 200 magnification

Fig. 6

Immunohistological staining of CD-68-positive cells (a) day 3, (b) day15, and (c) day 30 in the Creos™ Xenoprotect (CXP) group and (d) day 3, (e) day 15, and (f) day 30 in the control group. All graphs were captured at × 100 magnification. g Statistical analysis of the histomorphometrically measured CD-68-positive mononuclear cell number per square millimeter. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001