Sandra Reitmaier

Preliminary Investigations on Intradiscal Pressures during Daily Activities: An In Vivo Study Using the Merino Sheep

Purpose Currently, no studies exist, which attest the suitability of the ovine intervertebral disc as a biomechanical in vivo model for preclinical tests of new therapeutic strategies of the human disc. By measuring the intradiscal pressure in vivo, the current study attempts to characterize an essential biomechanical parameter to provide a more comprehensive physiological understanding of the ovine intervertebral disc. Methods Intradiscal pressure (IDP) was measured for 24 hours within the discs L2-L3 and L4-L5 via a piezo-resistive pressure sensor in one merino sheep. The data were divided into an activity and a recovery phase and the corresponding average pressures for both phases were determined. Additionally, IDPs for different static and dynamic activities were analyzed and juxtaposed to human data published previously. After sacrificing the sheep, the forces corresponding to the measured IDPs were examined ex vivo in an axial compression test. Results The temporal patterns of IDP where pressure decreased during activity and increased during rest were comparable between humans and sheep. However, large differences were observed for different dynamic activities such as standing up or walking. Here, IDPs averaged 3.73 MPa and 1.60 MPa respectively, approximately two to four times higher in the ovine disc compared to human. These IDPs correspond to lower ex vivo derived axial compressive forces for the ovine disc in comparison to the human disc. For activity and rest, average ovine forces were 130 N and 58 N, compared to human forces of 400-600 N and 100 N, respectively. Conclusions In vivo IDPs were found to be higher in the ovine than in the human disc. In contrast, axial forces derived ex vivo were markedly lower in comparison to humans. Both should be considered in future preclinical tests of intradiscal therapies using the sheep. The techniques used in the current study may serve as a protocol for measuring IDP in a variety of large animal models.

In vitro and in silico investigations of disc nucleus replacement

Currently, numerous hydrogels are under examination as potential nucleus replacements. The clinical success, however, depends on how well the mechanical function of the host structure is restored. This study aimed to evaluate the extent to and mechanisms by which surgery for nucleus replacements influence the mechanical behaviour of the disc. The effects of an annulus defect with and without nucleus replacement on disc height and nucleus pressure were measured using 24 ovine motion segments. The following cases were considered: intact; annulus incision repaired by suture and glue; annulus incision with removal and re-implantation of nucleus tissue repaired by suture and glue or plug. To identify the likely mechanisms observed in vitro, a finite-element model of a human disc (L4–L5) was employed. Both studies were subjected to physiological cycles of compression and recovery. A repaired annulus defect did not influence the disc behaviour in vitro, whereas additional nucleus removal and replacement substantially decreased disc stiffness and nucleus pressure. Model predictions demonstrated the substantial effects of reductions in replaced nucleus water content, bulk modulus and osmotic potential on disc height loss and pressure, similar to measurements. In these events, the compression load transfer in the disc markedly altered by substantially increasing the load on the annulus when compared with the nucleus. The success of hydrogels for nucleus replacements is not only dependent on the implant material itself but also on the restoration of the environment perturbed during surgery. The substantial effects on the disc response of disruptions owing to nucleus replacements can be simulated by reduced nucleus water content, elastic modulus and osmotic potential.

Is the ovine intervertebral disc a small human one? A finite element model study.

The sheep is one of the most frequently used animal models for experimental intervertebral disc research questions. Although there are large differences in size between human and ovine discs, recent in vivo and in vitro studies indicate similarities in the internal disc stresses. The present finite element model study, therefore, intended to detect the parameters that, despite the different geometry, ensure mechanical comparability between both species. At first, a finite element model of the human L4-L5 lumbar intervertebral disc was developed. The predicted displacement and nucleus pressure response were validated with experimental in vivo and in vitro data. Starting with adapting the model geometry from the human to the ovine disc, several material and biochemical parameters, which might contribute to the preservation of the mechanical disc response across both species, were successively adapted to ovine properties. Replacing the geometry yielded a substantially higher disc stiffness and lower nucleus pressure compared to in vitro measurements performed on ovine discs. Additional reduction of annulus and nucleus elasticity led to an improved correlation between model predictions and measurements. Changes in the glycosaminoglycan content and endplate permeability improved the predicted pressure, but only slightly affected the displacement response. Only the combination of all parameters resulted in a good agreement between the predictions and measurements. This study demonstrated that there are profound differences between model predictions and in vitro results if an ovine simulation is run with human material properties. However, once the species-specific material properties are included, the predictions fit the in vitro results. Therefore, it seems that the human and ovine disc is functionally adapted to produce similar internal stresses, despite the large variation in geometry.

Metaphyseal fracture healing in a sheep model of low turnover osteoporosis induced by hypothalamic–pituitary disconnection (HPD)

We recently established a large animal model of osteoporosis in sheep using hypothalamic–pituitary disconnection (HPD). As central regulation is important for bone metabolism, HPD‐sheep develop severe osteoporosis because of low bone turnover. In this study we investigated metaphyseal fracture healing in HPD‐sheep. To elucidate potential pathomechanisms, we included a treatment group receiving thyroxine T4 and 17β‐estradiol. Because clinically osteoporotic fractures often occur in the bone metaphysis, HPD‐sheep and healthy controls received an osteotomy in the distal femoral condyle. Half of the HPD‐sheep were systemically treated with thyroxine T4 and 17β‐estradiol during the healing period. Fracture healing was evaluated after 8 weeks using pQCT, µCT, and histomorphometrical analysis. Bone mineral density (BMD) and bone volume/total volume (BV/TV) were considerably reduced by 30% and 36%, respectively, in the osteotomy gap of the HPD‐sheep compared to healthy sheep. Histomorphometry also revealed a decreased amount of newly formed bone (−29%) and some remaining cartilage in the HPD‐group, suggesting that HPD disturbed fracture healing. Thyroxine T4 and 17β‐estradiol substitution considerably improved bone healing in the HPD‐sheep. Our results indicate that fracture healing requires central regulation and that thyroxine T4 and 17β‐estradiol contribute to the complex pathomechanisms of delayed metaphyseal bone healing in HPD‐sheep.

Increase or decrease in stability after nucleotomy? Conflicting in vitro and in vivo results in the sheep model

Nucleotomy is a common surgical procedure to treat disc herniations. The potential occurrence of segmental instability after surgery, however, is suspected to necessitate re-operation and fusion. Although in vitro studies support the theory of destabilization after nucleotomy, a prior, in-house animal study contrarily revealed an increase in stability after surgery. To identify which structural compartment of the motion segment is decisive for increased stability after nucleotomy in vivo, the flexibilities of ovine motion segments were measured after different stepwise reductions at the anterior and posterior spinal column. Different test groups were used in which nucleotomy had been performed during surgery in vivo and under isolated in vitro conditions, respectively. In accordance with expectations, in vitro nucleotomy on ovine motion segments significantly increased flexibility. By contrast, nucleotomy significantly decreased flexibility 12 weeks after surgery. After removal of the posterior structures, however, the differences in flexibility diminished. The present results thus suggest that it might not exclusively be the trauma to the intervertebral disc during surgery which is decisive for post-operative stability, but rather adaptive mechanisms in the posterior structures. Therefore, care should be taken to minimize the damage to the posterior structures in the course of the surgical approach, which more likely compromises stability.

Review of the fluid flow within intervertebral discs - How could in vitro measurements replicate in vivo?

By maintaining a balance between external mechanical loads and internal osmotic pressure, fluid content of intervertebral discs constantly alters causing fluctuations in disc hydration, height, diameter and pressure that govern disc temporal response. This paper reviews and discusses the relevant findings of earlier studies on the disc fluid flow with the aim to understand and remedy discrepancies between in vivo and in vitro observations. New results of finite element model studies are also exploited in order to help identify the likely causes for such differences and underlying mechanisms observed in vitro. In vivo measurements of changes in spinal height and disc fluid content/pressure via stadiometry, magnetic resonance imaging and intradiscal pressure measurements have been carried out. They have demonstrated that the disc volume, fluid content, height and nucleus pressure alter depending to a large extent on prior-current external load conditions. Although the diurnal loading lasts on average nearly twice longer than the subsequent resting (16 vs. 8h), the disc completely recovers its height and volume during the latter period through fluid inflow. In view of much longer periods required to recover disc height and pressure in vitro in ovine, porcine, caprine, bovine and rat discs, concerns have been raised on the fluid inflow through the endplates that might be hampered by clogged blood vessels post mortem. Analyses of discrepancies in the flow-dependent recoveries in vivo and in vitro highlight an excessive fluid content in the latter as a likely cause. To replicate in vivo conditions as closely as possible in vitro, preparation and preconditioning of specimens and/or pressure and osmolarity of the culture media in which specimens are immersed should hence be designed in a manner as to diminish disc hydration level and/or fluid transport.

Strontium(II) and mechanical loading additively augment bone formation in calcium phosphate scaffolds

Calcium phosphate cements (CPCs) are widely used for bone‐defect treatment. Current developments comprise the fabrication of porous scaffolds by three‐dimensional plotting and doting using biologically active substances, such as strontium. Strontium is known to increase osteoblast activity and simultaneously to decrease osteoclast resorption. This study investigated the short‐ and long‐term in vivo performances of strontium(II)‐doted CPC (SrCPC) scaffolds compared to non‐doted CPC scaffolds after implantation in unloaded or load‐bearing trabecular bone defects in sheep. After 6 weeks, both CPC and SrCPC scaffolds exhibited good biocompatibility and osseointegration. Fluorochrome labeling revealed that both scaffolds were penetrated by newly formed bone already after 4 weeks. Neither strontium doting nor mechanical loading significantly influenced early bone formation. In contrast, after 6 months, bone formation was significantly enhanced in SrCPC compared to CPC scaffolds. Energy dispersive X‐ray analysis demonstrated the release of strontium from the SrCPC into the bone. Strontium addition did not significantly influence material resorption or osteoclast formation. Mechanical loading significantly stimulated bone formation in both CPC and SrCPC scaffolds after 6 months without impairing scaffold integrity. The most bone was found in SrCPC scaffolds under load‐bearing conditions. Concluding, these results demonstrate that strontium doting and mechanical loading additively stimulated bone formation in CPC scaffolds and that the scaffolds exhibited mechanical stability under moderate load, implying good clinical suitability.

How do we stand? Variations during repeated standing phases of asymptomatic subjects and low back pain patients

An irreproducible standing posture can lead to mis-interpretation of radiological measurements, wrong diagnoses and possibly unnecessary treatment. This study aimed to evaluate the differences in lumbar lordosis and sacrum orientation in six repetitive upright standing postures of 353 asymptomatic subjects (including 332 non-athletes and 21 athletes - soccer players) and 83 low back pain (LBP) patients using a non-invasive back-shape measurement device. In the standing position, all investigated cohorts displayed a large inter-subject variability in sacrum orientation (∼40°) and lumbar lordosis (∼53°). In the asymptomatic cohort (non-athletes), 51% of the subjects showed variations in lumbar lordosis of 10-20% in six repeated standing phases and 29% showed variations of even more than 20%. In the sacrum orientation, 53% of all asymptomatic subjects revealed variations of >20% and 31% of even more than 30%. It can be concluded that standing is highly individual and poorly reproducible. The reproducibility was independent of age, gender, body height and weight. LBP patients and athletes showed a similar variability as the asymptomatic cohort. The number of standing phases performed showed no positive effect on the reproducibility. Therefore, the variability in standing is not predictable but random, and thus does not reflect an individual specific behavioral pattern which can be reduced, for example, by repeated standing phases.

Spinal fusion without instrumentation – Experimental animal study

Background The number and cost of instrumented spinal fusion surgeries have increased rapidly, primarily for the treatment of lumbar segmental instabilities. However, what if the organism itself is able to restore segmental stability over time? This large‐animal study using sheep aimed to investigate whether the reparative response after destabilization via facetectomy and nucleotomy without instrumentation can effectively fuse the spinal segment comparable to instrumented standard fusion surgery. Methods The following four surgical interventions were investigated: dorsal fixation via internal fixator, ventral fixation via cage as well as facetectomy and nucleotomy without additional instrumentation. Six months postoperatively, the animals were sacrificed, and the lumbar spines were used for biomechanical tests. Findings Spinal stability was restored to the destabilized spinal segments at six months postoperatively and was comparable to the results of conventional surgery via screws and cages. Iatrogenic hypomobilization caused significant reductions in facet joint space and intervertebral disc height of segments at index and adjacent level. Restabilized segments after iatrogenic hypermobilzation also significantly decreased facet joint space and disc height at index level, but revealed no influence on adjacent segments. Interpretation These findings in the sheep model question the necessity of costly instrumentation and suggest the alternative possibility of stimulating the reparative capacity of the body in human lumbar spine fusion surgery. HighlightsAnimal study on fusion after spinal destabilization with and without instrumentationSpinal stability was restored six months postoperatively without instrumentation.Results were comparable to conventional surgery via screws and cages.Findings challenge the necessity of costly instrumentation.

Intramembranous bone formation after callus distraction is augmented by increasing axial compressive strain

The mechanical environment is a primary factor in the success of distraction osteogenesis. It is known that the interfragmentary movement during the distraction and maturation phase effects the callus formation. In addition to cyclic compression, other movements like shear and bending influence the bone formation process as shown in previous callus distraction studies. Reports of cartilage presence and endochondral ossification in the regenerative zone have been associated with a lack of fixation stability and delayed healing. So far the effects of the direction of interfragmentary movements could not be studied separately. By means of a unique lateral callus distraction model, we investigated the effects of small (0.1 mm) and moderate (0.6 mm), purely axial compression on ossification during callus maturation in sheep. A distraction device incorporating a mobile titanium plate was mounted on the tibia. Following lateral callus distraction, electromechanically controlled movements allowed purely axial cyclic compression of the tissue regenerate. Seven weeks post-operatively, the tissue regenerates were investigated using μCT, histology and immunohistochemistry. The larger amplitude significantly increased bone formation (Fractional bone volume: 19.4% vs. 5.2%, p = 0.03; trabecular thickness: 0.1 mm vs. 0.06 mm, p = 0.006; mean spicule height: 2.6 mm vs. 1.1 mm, p = 0.02) however, no endochondral ossification occurred. The elimination of shear movement, unimpaired neovascularization as well as the tensile strain stimuli during the distraction phase suppressing chondrogenic differentiation may all contribute to the absence of cartilage. In clinical application of distraction osteogenesis, moderate axial interfragmentary movement augments intramembranous ossification provided shear strain is minimized.