Karl H. Wenger

Biomechanical evaluation of the stability of thoracolumbar burst fractures.

Study Design. The decision to treat thoracolumbar burst fractures in neurologically intact patients either surgically or nonoperatively depends largely on whether the fracture is clinically stable. This study evaluated the relative contributions of the anterior, middle, and posterior columns to spinal stability by way of in vitro experimentation and supplemental analysis of patients with nonoperatively treated burst fractures. Methods. An L1 burst fracture model was used to evaluate the contribution of the three columns of the spine to resisting imposed flexion deforming forces. Six spines were tested to a gross bending flexion angle of 25°. Changes in vertebral motion across the site of injury were measured and compared. In addition, a summary of our recent clinical experience with nonoperatively treated burst fractures is presented and correlated with the studys laboratory findings. Results. T12–L2 motion measurements after vertebral and ligamentous disruption revealed a statistically significant increase in motion upon anterior and added posterior column compromise, but not for added middle column disruption. Review of the clinical series revealed that burst fractures with anterior and middle column compromise but an intact posterior column were stable and healed satisfactorily. Conclusions. The data suggest that the condition of the posterior column, not the middle column, is a better indicator of burst fracture stability. It is proposed that the classic burst fracture (anterior and middle column compromise) is a stable injury that, in the absence of neurologic deficit, can be managed nonoperatively.

Age-Related Changes in the Osteogenic Differentiation Potential of Mouse Bone Marrow Stromal Cells

Age‐dependent bone loss has been well documented in both human and animal models. Although the underlying causal mechanisms are probably multifactorial, it has been hypothesized that alterations in progenitor cell number or function are important. Little is known regarding the properties of bone marrow stromal cells (BMSCs) or bone progenitor cells during the aging process, so the question of whether aging alters BMSC/progenitor osteogenic differentiation remains unanswered. In this study, we examined age‐dependent changes in bone marrow progenitor cell number and differentiation potential between mature (3 and 6 mo old), middle‐aged (12 and 18 mo old), and aged (24 mo old) C57BL/6 mice. BMSCs or progenitors were isolated from five age groups of C57BL/6 mice using negative immunodepletion and positive immunoselection approaches. The osteogenic differentiation potential of multipotent BMSCs was determined using standard osteogenic differentiation procedures. Our results show that both BMSC/progenitor number and differentiation potential increase between the ages of 3 and 18 mo and decrease rapidly thereafter with advancing age. These results are consistent with the changes of the mRNA levels of osteoblast lineage‐associated genes. Our data suggest that the decline in BMSC number and osteogenic differentiation capacity are important factors contributing to age‐related bone loss.

Lower Uncarboxylated Osteocalcin Concentrations in Children with Prediabetes Is Associated with β-Cell Function

CONTEXT Although animal studies suggest that it is the uncarboxylated rather than carboxylated form of osteocalcin that affects glucose homeostasis, the human data are scant and equivocal. OBJECTIVE This study investigated associations of uncarboxylated and carboxylated forms of osteocalcin with insulin sensitivity and β-cell function in 140 overweight prepubertal children (43% female, 46% black, 84% obese) with normal glucose levels (n = 99) and prediabetes (n = 41). METHODS An oral glucose tolerance test was used to identify prediabetes and for measurement of insulin sensitivity (Matsuda index), β-cell function [oral glucose tolerance test derived insulinogenic index and disposition index (DI(OGTT))] and uncarboxylated and carboxylated forms of osteocalcin. Visceral adipose tissue (VAT) was assessed using magnetic resonance imaging. RESULTS After controlling for age, sex and race, lower uncarboxylated osteocalcin concentrations, Matsuda index, insulinogenic index, and DI(OGTT) and higher VAT levels were found in the prediabetes vs. normal-glucose group (all P < 0.03). Carboxylated osteocalcin levels were not different between groups. Multiple linear regression adjusting for age, sex, race, and VAT revealed that uncarboxylated osteocalcin was associated with insulinogenic index and DI(OGTT) (β = 0.34, 0.36, respectively, both P < 0.04) in the prediabetes group but not the normal-glucose group. In both the normal-glucose and prediabetes groups, carboxylated osteocalcin was associated with insulin sensitivity (β = 0.26, 0.47, respectively, both P < 0.02). CONCLUSIONS These data suggest that the lower uncarboxylated osteocalcin concentrations found in children with prediabetes may be associated with β-cell dysfunction. In addition, our findings between carboxylated osteocalcin and insulin sensitivity suggest that carboxylated osteocalcin plays a role in human glucose homeostasis.

Myostatin (GDF-8) Deficiency Increases Fracture Callus Size, Sox-5 Expression, and Callus Bone Volume

Myostatin (GDF-8) is a negative regulator of skeletal muscle growth and mice lacking myostatin show increased muscle mass. We have previously shown that myostatin deficiency increases bone strength and biomineralization throughout the skeleton, and others have demonstrated that myostatin is expressed during the earliest phase of fracture repair. In order to determine the role of myostatin in fracture callus morphogenesis, we studied fracture healing in mice lacking myostatin. Adult wild-type mice (+/+), mice heterozygous for the myostatin mutation (+/-), and mice homozygous for the disrupted myostatin sequence (-/-) were included for study at two and four weeks following osteotomy of the fibula. Expression of Sox-5 and BMP-2 were significantly upregulated in the fracture callus of myostatin-deficient (-/-) mice compared to wild-type (+/+) mice at two weeks following osteotomy. Fracture callus size was significantly increased in mice lacking myostatin at both two and four weeks following osteotomy, and total osseous tissue area and callus strength in three-point bending were significantly greater in myostatin -/- mice compared to myostatin +/+ mice at four weeks post-osteotomy. Our data suggest that myostatin functions to regulate fracture callus size by inhibiting the recruitment and proliferation of progenitor cells in the fracture blastema. Myostatin deficiency increases blastema size during the early inflammatory phase of fracture repair, ultimately producing an ossified callus having greater bone volume and greater callus strength. While myostatin is most well known for its effects on muscle development, it is also clear that myostatin plays a significant, direct role in bone formation and regeneration.

Effect of whole-body vibration on bone properties in aging mice

Recent studies suggest that whole-body vibration (WBV) can improve measures of bone health for certain clinical conditions and ages. In the elderly, there also is particular interest in assessing the ability of physical interventions such as WBV to improve coordination, strength, and movement speed, which help prevent falls and fractures and maintain ambulation for independent living. The current study evaluated the efficacy of WBV in an aging mouse model. Two levels of vibration--0.5 and 1.5g--were applied at 32Hz to CB57BL/6 male mice (n=9 each) beginning at age 18 months and continuing for 12 weeks, 30 min/day, in a novel pivoting vibration device. Previous reports indicate that bone parameters in these mice begin to decrease substantially at 18 months, equivalent to mid-fifties for humans. Micro-computed tomography (micro-CT) and biomechanical assessments were made in the femur, radius, and lumbar vertebra to determine the effect of these WBV magnitudes and durations in the aging model. Sera also were collected for analysis of bone formation and breakdown markers. Mineralizing surface and cell counts were determined histologically. Bone volume in four regions of the femur did not change significantly, but there was a consistent shift toward higher mean density in the bone density spectrum (BDS), with the two vibration levels producing similar results. This new parameter represents an integral of the conventional density histogram. The amount of high density bone statistically improved in the head, neck, and diaphysis. Biomechanically, there was a trend toward greater stiffness in the 1.5 g group (p=0.139 vs. controls in the radius), and no change in strength. In the lumbar spine, no differences were seen due to vibration. Both vibration groups significantly reduced pyridinoline crosslinks, a collagen breakdown marker. They also significantly increased dynamic mineralization, MS/BS. Furthermore, osteoclasts were most numerous in the 1.5 g group (p≤ 0.05). These findings suggest that some benefits of WBV found in previous studies of young and mature rodent models may extend to an aging population. Density parameters indicated 0.5 g was more effective than 1.5 g. Serological markers, by contrast, favored 1.5 g, while biomechanically and histologically the results were mixed. Although the purported anabolic effect of WBV on bone homeostasis may depend on location and the parameter of interest, this emerging therapy at a minimum does not appear to compromise bone health by the measures studied here.

Periodontal ligament fibroblasts sustain destructive immune modulators of chronic periodontitis.

BACKGROUND In healthy periodontal tissue, innate immune responses effectively confine and suppress a bacterial insult. However, a disruption of the host-bacterial equilibrium may produce an overexpression of cytokines and lead to permanent, host-mediated tissue damage. Although such periodontal destruction primarily results from activated immune mechanisms, the site-specific damage suggests that local tissues participate in these pathologic changes. Periodontal ligament fibroblasts (PDLFs) are prominent in the periodontium and are critical in homeostasis and regeneration because they have the ability to produce multiple cytokines in response to a bacterial insult. These cells could play a role in the local pathogenesis of periodontal disease. METHODS We studied alkaline phosphatase (ALP) activity, interleukin (IL)-6 production, and morphologic characteristics of cultured PDLFs that were isolated from periodontally healthy sites (H-PDLFs) and diseased sites (D-PDLFs) in humans. Quantitative analyses of 84 genes that are related to inflammation were performed using real-time polymerase chain reaction arrays. RESULTS A mineralizing medium induced a significant increase of ALP in H-PDLFs, but no significant enzymatic changes were detected in D-PDLFs after such treatment. The protein and gene expression of IL6 showed a significant upregulation in D-PDLFs, which also demonstrated a significant upregulation of 54% of genes in the inflammatory gene arrays. CONCLUSIONS To our knowledge, these results represent the first biologic evidence that D-PDLFs retain uniquely inflammatory phenotypes that could maintain localized destructive signals in periodontitis. The overexpression of proinflammatory cytokines by PDLFs could amplify local inflammation by the continuous triggering of immune responses. In addition, the location of these cells could be critical in the progression of the inflammatory front into the deeper tissues.

Matrix remodeling expression in anulus cells subjected to increased compressive load.

Study Design. Mechanobiology study of gene expression changes as a result of compressive overload of anular fibrochondrocytes. Objective. To test hypotheses regarding phenotype shift in genes coding for representative extracellular matrix (ECM) proteins and matrix modulators. Summary of the Background Data. In degenerative disc disease, the transfer of compressive load through the disc shifts largely from the nucleus onto the anulus. In vivo models simulating this condition have shown derangement of the collagenous ultrastructure in the anulus. In vitro models of cultured anulus cells subjected to static compressive stress generally suggest a down-regulation of synthesis. This study evaluated the expression of specific isomers of genes responsible for mechanical viability and metabolism of the disc under cyclic compressive loads. Methods. Fibrochondrocytes were digested from the anuli of 3, 2-week-old pigs, embedded in 1.5% alginate gel, and hydrostatically compressed at 0.5 Hz for 3 hours to amplitudes of 10 and 30 atm. These levels represented nominal load transfer through the healthy disc and high load transfer through the degenerative disc. Ribonucleic acid was isolated, reverse transcribed, and evaluated by real-time polymerase chain reaction for expression of type I (C-I) and type II (C-II) collagen, aggrecan, the matrix metalloproteinase (MMP-1), and the transforming growth factor beta (TGF&bgr;-1). Results were expressed at percentages of uncompressed controls. Results. The lower pressure of 10 atm resulted in up-regulation of all ECM protein genes. C-I and C-II both averaged 141%, and aggrecan 121% of controls (P < 0.05). MMP-1 and TGF&bgr;-1 were essentially unchanged. With the pressure increased to 30 atm, C-II remained approximately at the level expressed under lower pressure, but C-I was reduced to 42% of controls (P < 0.05), indicating a phenotype shift. MMP-1 and TGF&bgr;-1 also were down-regulated to 71% and 54% of controls, respectively (P < 0.05). Conclusions. The up-regulation of the ECM genes with nominal pressure highlights the mechanobiological importance of common activity in fibrocartilage homeostasis. Differential regulation of the 2 primary collagen types with high pressure indicates a capacity of the anulus to remodel according to pathomechanical conditions.

Effects of facetectomy and crosslink augmentation on motion segment flexibility in posterior lumbar interbody fusion.

STUDY DESIGN Biomechanical assessment using calf lumbar motion segments. OBJECTIVE To determine whether facetectomy affects the primary stability of posterior lumbar interbody fusion. SUMMARY OF BACKGROUND DATA To improve visualization and access to the disc space, the facet joints frequently are removed. Previous biomechanical studies have indicated a fundamental role for the facet joints in maintaining spinal segment stability. METHODS Single motion segments from calf lumbar spines were tested for pure-moment flexibility in flexion-extension (FE), lateral bending (LB), and axial rotation (AR). After testing intact, an interbody cage and pedicle screw system were implanted. Next, a bilateral facetectomy was performed, and finally a crosslink was added. Flexibility testing was repeated at each stage of implantation. Data are reported for range of motion (ROM), neutral zone (NZ), and a new compliance parameter (COM), based on the slopes of the moment-angle curve in the neutral and elastic regions. RESULTS With posterior lumbar interbody fusion implantation, ROM in FE was reduced 82% +/- 4% (mean +/- standard deviation) and NZ 78% +/- 7% over intact (P < 0.015: Wilcoxon). Reduction in LB was slightly more, whereas reduction in AR was considerably less and did not achieve statistical significance for NZ. After facetectomy, ROM in FE increased 0.3 degrees (P < 0.05), on average, and NZ did not change. In LB neither changed significantly. In AR, ROM increased 0.6 degrees (P < 0.05), and NZ increased 0.2 degrees (P < 0.05). The addition of a crosslink changed ROM and NZ less than 0.1 degrees in FE and LB, whereas in AR it restored half of the stability lost due to facetectomy in ROM (P < 0.05), and had a similar trendwise effect on NZ. The new compliance measure, COM, was found to agree with the direction of change in ROM more consistently than did NZ. CONCLUSION Facetectomy causes a nominal increase in ROM and NZ in FE and LB, which are not affected by the addition of a crosslink. Although the effect of facetectomy is greater in AR-and crosslink has a measurable restoring effect-all differences are within a few tenths of a degree under this loading paradigm. Thus, the clinical utility of adding a crosslink may not be justified based on these small biomechanical changes. COM can serve as a complement to ROM and NZ, or even as a surrogate when its 2 components are reported together, as it shows strong agreement with ROM, effectively distinguishes between lax and elastic region behaviors, and provides a measure of flexibility independent of the load range.Study Design. In vitro testing of vertebroplasty techniques including pulsed jet-lavage for fat and marrow removal in human cadaveric lumbar and thoracic vertebrae. Objective. To develop jet-lavage techniques for vertebroplasty and investigate their effect on cement distribution, injection forces, and fat embolism. Summary of Background Data. The main complications of cement vertebroplasty are cement leakage and pulmonary fat embolism, which can have fatal consequences and are difficult to prevent reliably by current vertebroplasty techniques. Methods. Twenty-four vertebrae (Th8–L04) from 5 osteoporotic cadaver spines were grouped in triplets depending on bone mineral density (BMD). Before polymethylmethacrylate (PMMA) vertebroplasty, a pulsatile jet-lavage for removal of intertrabecular fat and bone marrow was performed in 2 groups with 8 specimens each, performing radial and axial irrigation from the biopsy needles. One hundred mL of Ringer solution were injected through 1 pedicle and regained by low vacuum via the contralateral pedicle. Eight control vertebrae were not irrigated. All specimens underwent standardized PMMA cement augmentation injecting 20% of the vertebral volume. Injection forces, cement distribution, and extravasations were quantified. Results. All irrigation solution could be retrieved with the vacuum applied. A Kruskal-Wallis test revealed significantly higher injection forces of the control group as compared with the irrigated groups (P = 0.021). Dilatation of the syringe at forces above 300 N occurred in 75% of the untreated compared with 12.5% of the lavaged specimens. CT distribution analysis showed more homogenous cement distribution of the cement and significantly less extravasation in the irrigated specimens. Conclusion. The developed lavage technique for vertebroplasty showed to be feasible and reproducible. The reduction of injection forces would allow the use of more viscous PMMA cement lowering the risk for cement embolization and results in a safer procedure. The wash-out of bone marrow and the possible reduction of pulmonary fat embolism have to be verified with in vivo models.

Role of Myostatin (GDF-8) Signaling in the Human Anterior Cruciate Ligament

Myostatin, also referred to as growth and differentiation factor‐8 (GDF‐8), is expressed in muscle tissue where it functions to suppress myoblast proliferation and myofiber hypertrophy. Recently, myostatin and its receptor, the type IIB activin receptor (ActRIIB), were detected in the leg tendons of mice, and recombinant myostatin was shown to increase cellular proliferation and the expression of type 1 collagen in primary fibroblasts from mouse tendons. We sought to determine whether myostatin and its receptor were present in human anterior cruciate ligament (ACL) tissue, and if myostatin treatment had any effect on primary ACL fibroblasts. ACL tissue samples were obtained from material discarded during ACL reconstruction surgery. Real‐time PCR and immunohistochemistry demonstrate that both myostatin and its receptor are abundant in the human ACL. Primary fibroblasts isolated from human ACL specimens were treated with recombinant myostatin, and myostatin treatment increased fibroblast proliferation as well as the expression of tenascin C (TNC), type 1 collagen, and transforming growth factor‐β1. Real‐time PCR analysis of TNC and type 1 collagen expression in ACL specimens from normal mice and mice lacking myostatin supported these findings by showing that both TNC and type 1 collagen were downregulated in ACL tissue from myostatin‐deficient mice. Together, these data suggest that myostatin is a pro‐fibrogenic factor that enhances cellular proliferation and extracellular matrix synthesis by ACL fibroblasts. Recombinant myostatin may therefore have therapeutic applications in the area of tendon and ligament engineering and regeneration.

New technology applications: Knotless barbed suture for tracheal resection anastomosis.

Tracheal resection anastomoses are often under tension and can be technically challenging. New suture materials such as V‐loc (barbed, knotless wound closure device) may offer advantages over conventional methods. The objective of this study is to determine if a running V‐loc suture is of comparable tensile strength to conventional closure.