OTA 1997 Posters - Scientific Basis for Fracture Care

*Distraction Osteogenesis in a Micropig Animal Model. An Alternative Sequence of Regenerate Formation

Hermann Josef Bail, MD, M. Raschke, S. Kolbeck, A. Weiler, N. P. Haas

Humboldt University, Berlin, Germany

Purpose: Distraction osteogenesis is a widely accepted method for limb lengthening and treatment of bone defects caused by trauma, tumor or infection. Based on animal models and biopsies from human distraction regenerates, it was hypothesized that bone formation is achieved exclusively through intramembranous ossification (Ilizarov 1994). According to this theory, newly formed bone is deposited on collagen fibers that span the bone ends and are oriented in the direction of distraction (Aronson 1990). In this study, we sought to determine whether a histological examination of a porcine model of distraction osteogenesis would support this theory of bone formation.

Methods: The left tibiae and fibulae of micropigs were osteotomized with an oscillating saw, opening the periosteum before and sewing it again after this procedure. Using an external half ring fixator, the tibiae were stabilized and distracted. The pigs were divided into three groups with different modalities of distraction and consolidation (Tab. 1). Each group received in-vivo labeling with calcein-green during the latency period, tetracycline at the end of distraction and xylenolorange at the end of consolidation. Distraction regenerates including the neighboring cortical bone, were harvested after sacrifice. Non-decalcified sagitally orientated 6µm slices were stained with astra-blue, safranin-O, according to the von-Kossa method and with Masson-Goldners trichrome stain and then analyzed with light and polarized light microscopy. 80µm-thick grinds of the same orientation were investigated with fluorescence light microscopy.

Tab. 1

	group 1 (n=5)	group 2 (n=4)	group 3 (n=5)
distraction mode	2 mm/day	1.5 mm/day	1 mm/day
latency period	4 days	4 days	4 days
distraction period	10 days	10 days	10 days
consolidation period	11 days	16 days	26 days

Results: The regenerates of group 3 were completely consolidated; newly formed callus was bridging the distraction gap and the bone ends had a partially loosened structure, indicating remodeling. In contrast, group 2 showed connection of the callus ends in only some areas, while in group 1, a fibrous interzone in the middle of the gap was always detectable. Islands of chondral ossification were observed in the regenerates of group 2 and 1. Polarized light microscopy of the fibrous interzone showed parallel collagen fibers oriented according to the distraction direction in most cases. The observed chondrocytes were located preferably in a crisscross collagen fiber environment. Fluorescence microscopy showed that callus formation started from periosteum and sometimes from endosteum in each of the groups.

Discussion: Group 3 showed bony consolidation of the regenerate, demonstrating that distraction can be successfully carried out in micropigs using our model. Cartilage islands occurring within the criss-cross oriented collagen fibers in groups 1 and 2 indicate temporary mechanical instability in the regenerate. Ilizarov (1984) and Aronson (1990) described collagen fiber formation as derived from the cortical bone ends; however, these studies involved a corticotomy, after which the medullar artery was left intact. May be, if there is any damage to the medullar artery, the surface of the cortical bone does not provide an environment for the formation of collagen tissue. In this case, periosteal and endosteal formatted callus seems to function as a host bone surface for the collagen membranes.

Conclusion: Our investigation indicates that a distractible callus can be also formatted by the periosteum and endosteum alone. This may demonstrate the importance of periostal restoration when the medullar canal is transversely sectioned during surgical procedure.

References

Aronson-J; Good-B; Stewart-C; Harrison-B; Harp-J: Preliminary studies of mineralization during distraction osteogenesis; Clin-Orthop. 1990 Jan(250): 43-9 llizarov-GA; Barabash-AP; Imerlishvili-AA; Larionov-AA; Kochetkov-luS: Morphological characteristics of the formation and reconstruction of bone tissue in the replacement of extensive bone defects; OrtopTravmatol-Protez. 1984 Jan(l): 16-20.