Clinical StudiesPosterolateral lumbar fusions in athymic rats: characterization of a model☆
Introduction
Spinal fusion is a common surgical procedure, and autograft is the gold standard grafting material. However, despite advances in surgical techniques, pseudarthrosis remains an issue [1]. Further, autograft may be limited in supply, and its harvest is associated with significant morbidity and increased operative time [2]. Attention has thus been directed toward refining alternative means of inducing bone formation.
Bone graft substitutes that are currently being studied include demineralized bone matrices (DBMs), individual recombinant bone morphogenetic proteins (BMPs) and autogenous blood product isolates. In addition to the active agent in these new products, the mode of delivery and type of carrier are important factors in determining efficacy.
With the many variables being studied, it has been necessary to use animal models. Noninstrumented posterolateral lumbar fusions are commonly studied, because they pose a challenging and clinically relevant model. Studies have been done in a number of species, including rats [3], [4], rabbits [5], [6] and dogs [7], [8]. Products are often tested in lower species and moved along to higher species only when promising results are found.
Some osteoinductive products, however, cannot be tested in certain models. For example, Aspenburg et al. [9] found xenogenic DBM to induce little or no bone formation when placed intramuscularly in normal rats. It was hypothesized that this was because of an immunologic response of the host to the nonconserved or differentially expressed donor proteins. Substantiating this hypothesis, increased bone induction was seen when athymic recipient rats were studied under otherwise identical circumstances. Although xenogenic DBMs are not used clinically, there is interest in studying them in animals as products are being developed. Other studies have shown human DBM to have a dose-dependent osteoinductive effect when placed intramuscularly or subcutaneously in athymic rats [10].
Additional work is ongoing to study the effectiveness of BMP containing adenoviral vectors (Ad) in inducing bone formation. When Ad-BMP-2 was injected intramuscularly into normothymic rats, significant immune response (without concomitant bone formation) was elicited by the first-generation adenoviral constructs [11], [12]. Conversely, bone formation without such immune response was seen when the same construct was studied in athymic rats.
Subsequently, the athymic rat has been used to study the potential role of osteoinductive products in spinal fusions by several authors. Athymic rat fusions have been studied with open implantation of xenogenic DBMs [4], [13], injection of adenoviral constructs [14], [15] and application of mammalian expression vectors [3].
Much evidence suggests that nonsteroidal anti-inflammatory agents can adversely affect fusion rates [16], [17]. Should a similar effect be expected if fusions are studied in the athymic rat where the immune system is compromised? Contrary to what might be expected, bone formation and metabolism does not appear to be affected by the absence of the thymus gland in rats [18]. Another study showed no significant differences in the healing of osteotomized tibias of athymic and normothymic rats [19].
Some small animal models have high fusion rates solely because of surgical exposure. This is not expected in the athymic rat model. Entire study groups have been described by Wang et al. [4] with no fusion (namely with Dynagraft [Citagenix, Inc., Lavai, QC, Canada], which was thought to be of insufficient osteoinductivity). By inference, fusion does not appear to be induced by surgical approach alone. This, however, has not yet been demonstrated as an independent variable.
Although used in previously reported studies, the model for open posterolateral lumbar fusion in the athymic rat has not, to our knowledge, been well characterized as performed for other fusion models [5]. The purpose of the current study is to clearly characterize a posterolateral lumbar fusion model in the athymic rat. Radiographic, manual palpation and histologic examinations are to be used to define the rates of autograft fusion, effect of an absent thymus and rates of spontaneous fusion in this model.
Section snippets
Overview
Sixty L4–L5 posterolateral lumbar fusions were performed (of note, rats have six lumbar vertebrae). Forty were mature female athymic nude rats (rnu/rnu), 8 to 9 weeks of age. Twenty received autograft (experimental study group, athymic/autograft). Twenty were opened and closed with no graft placed (negative control group, athymic/no graft). An additional 20 fusions were performed on 9- to 10-week-old immunologically intact female Sprague Dawley rats. These animals were similar to the athymic
Results
The athymic rats used in this experiment weighed between 170 and 200 g. The normothymic rats weighted between 190 and 200 g. The surgical procedures took approximately 20 to 30 minutes to perform.
Three rats were excluded from the study because of complications: two were lost perioperatively because of anesthesia, and one was lost approximately 1 week postoperatively with no clear cause. Of the three rats lost, all were normothymic. These three rats were replaced and are not included in the
Discussion
The athymic rat model has previously been used to study spinal fusion, but additional characterization was required before continuing study with this useful model. In what frame of reference should posterolateral fusion studies performed in these animals be interpreted?
The rat is a small and easily handled animal in which posterolateral fusion is easy to perform. Although much smaller than the human, the morphology of the rat lumbar vertebrae are roughly similar to that of the human (Fig. 3).
Acknowledgements
The authors thank Daniel Zelterman, PhD, for his statistical assistance and Chris Coady for her histologic assistance.
References (21)
- et al.
Osseointegration of autograft versus osteogenic protein-1 in posterolateral spinal arthrodesis: emphasis on the comparative mechanisms of bone induction
Spine J
(2002) - et al.
Pseudarthosis of the spine
Clin Orthop
(1992) - et al.
Perioperative and long-term complications of iliac crest bone graft harvesting for spinal surgery: a quantitative review of the literature
Int Med J
(2001) - et al.
Lumbar spine fusion by local gene therapy with cDNA encoding a novel osteoinductive protein (LMP-1)
Spine
(1998) - Wang JC, Davies M, Kanim LEA, Ukatu CJ, Dawson EG, Lieberman JR. Prospective comparison of commercially available...
- et al.
An experimental lumbar intertransverse process spinal fusion model: radiographic, histologic, and biomechanical healing characteristics
Spine
(1995) - et al.
Evaluation of OP-1 as a graft substitute for intertransverse process lumbar fusion
Spine
(2001) - et al.
Lumbar spinal fusion using recombinant human bone morphogenetic protein in the canine: a comparison of three dosages and two carriers
Spine
(1999) - et al.
Rabbit bone matrix induces bone formation in the athymic rat
Acta Orthop Scand
(1988) - et al.
Osteoinduction of human demineralized bone: characterization in a rat model
Clon Orthop Rel Res
(1998)
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FDA device/drug status: not applicable.
Author JNG acknowledges a financial relationship (grant research support from Stryker Biotech, Hopkinton, MA), which may indirectly relate to the subject of this research. Additional support was received from the Yale Core Center for Musculoskeletal Disorders.