Molly T. Vogt

Modulation of the biologic activity of the rabbit intervertebral disc by gene therapy: an in vivo study of adenovirus-mediated transfer of the human transforming growth factor beta 1 encoding gene.

Study Design. In vivo studies using a rabbit model to determine the biologic effects of direct, adenovirus-mediated transfer of a therapeutic gene to the intervertebral disc.Objectives.1) To deliver an exogenous therapeutic gene to rabbit lumbar intervertebral discs in vivo, 2) to quantify the resulSTUDY DESIGN In vivo studies using a rabbit model to determine the biologic effects of direct, adenovirus-mediated transfer of a therapeutic gene to the intervertebral disc. OBJECTIVES 1) To deliver an exogenous therapeutic gene to rabbit lumbar intervertebral discs in vivo, 2) to quantify the resulting amount of gene expression, and 3) to determine the effect on the biologic activity of the discs. SUMMARY OF BACKGROUND DATA Although growth factors such as transforming growth factor beta 1 appear to have promising therapeutic properties, there currently is no practical method for sustained delivery of exogenous growth factors to the disc for the management of certain chronic types of disease (e.g., disc degeneration). A possible solution is to modify the disc cells genetically through gene transfer such that the cells manufacture the desired growth factors endogenously on a continuous basis. METHODS Saline, with or without virus, was injected directly into lumbar discs of 22 skeletally mature female New Zealand white rabbits. Group 1 (n = 11) received the adenovirus construct Ad/CMV-hTGF beta 1 containing the therapeutic human transforming growth factor beta 1-encoding gene. Group 2 (n = 6) received adenovirus containing the luciferase marker gene. Group 3 (n = 5) received saline only. The rabbits were killed 1 week after injection. Immunohistochemical staining for human transforming growth factor beta 1 was performed on the disc tissues of one rabbit from Group 1. Nucleus pulposus tissues from the remaining rabbits were cultured in serumless medium. Bioassays were performed to determine human transforming growth factor beta 1 production and proteoglycan synthesis. RESULTS Discs injected with Ad/CMV-hTGF beta 1 exhibited extensive and intense positive immunostaining for transforming growth factor beta 1. The nucleus pulposus tissues from the discs injected with Ad/CMV-hTGF beta 1 exhibited a 30-fold increase in active transforming growth factor beta 1 production, and a 5-fold increase in total (active + latent) transforming growth factor beta 1 production over that from intact control discs (P < 0.05). Furthermore, these tissues exhibited a 100% increase in proteoglycan synthesis compared with intact control tissue, which was statistically significant (P < 0.05). CONCLUSIONS The results of this study suggest that the intervertebral disc is an appropriate site for adenovirus-mediated transfer of exogenous genes and subsequent production of therapeutic growth factors. Gene therapy therefore may have useful applications for study of the basic science of the intervertebral disc and for clinical management of degenerative disc disease.

Lower extremity arterial disease and the aging process: A review

Atherosclerotic disease is a major health problem in the elderly population in the U.S. To date, most research has focused on the coronary and cerebral manifestations of the disease. Lower extremity arterial disease (LEAD) has received less attention. However, the use of simple objective non-invasive diagnostic techniques in recent population-based studies has allowed identification of asymptomatic LEAD and shown that the prevalence of disease is several fold higher than previously estimated. The prevalence increases sharply with age, from 3% in those under 60 years to over 20% at 75+ years. Detection of LEAD in the elderly provides a rapid, easy assessment of the level of atherosclerosis in the body and identifies those at high risk for mortality and morbidity. LEAD is associated with a relative risk of 4 to 5 for all cause mortality. In about 25% of those with LEAD, the disease progresses over time, leading to loss of mobility, gangrene or amputation. This review summarizes current knowledge regarding the etiology and natural history of LEAD from an epidemiologic viewpoint, delineating areas in which additional research is needed.

A slowly progressive and reproducible animal model of intervertebral disc degeneration characterized by MRI, X-ray, and histology

Study Design. The progression of intervertebral disc degeneration following anterolateral “stab” of adult rabbit lumbar discs by 16-gauge hypodermic needle to a limited (5-mm) depth was studied for up to 24 weeks using magnetic resonance imaging, radiograph, and histologic outcome measures. Objectives. To develop a slowly progressive, reproducible rabbit model of intervertebral disc degeneration suitable for studying pathogenesis and pathophysiology of intervertebral disc degeneration and testing safety and efficacy of novel approaches to the treatment of intervertebral disc degeneration (e.g., growth factors, gene therapy, cell therapy, and tissue engineering). Summary of Background Data. Numerous animal models of intervertebral disc degeneration have been proposed in the literature, each with attendant advantages and disadvantages. The classic “stab model,” involving full-thickness stab of anterior anulus fibrosus of adult rabbit lumbar discs by a number 11 scalpel blade, appears to produce changes in certain biochemical and histologic outcome measures that are similar to changes seen in human intervertebral disc degeneration. However, the immediate herniation of nucleus pulposus on full-thickness stab renders this model less suitable for 1) studying effects of less precipitous changes in nucleus pulposus and anulus fibrosus that may be important in the onset and progression of intervertebral disc degeneration and 2) testing novel therapeutic approaches that target the processes of early intervertebral disc degeneration. Methods. The L2–L3, L3–L4, and L4–L5 lumbar intervertebral discs of 18 skeletally mature female New Zealand White rabbits were stabbed by 16-gauge hypodermic needle to a depth of 5 mm in the left anterolateral anulus fibrosus. Serial magnetic resonance imaging scans of the stabbed discs and intact L1–L2 and L5–L6 control discs were performed at 3, 6, 12, and 24 weeks post surgery and compared with preoperative magnetic resonance images. Supplemental radiograph and histologic analyses were performed. Results. The stabbed discs exhibited a progressive decrease in “magnetic resonance imaging index” (the product of nucleus pulposus area and signal intensity from T2-weighted midsagittal plane images) starting at 3 weeks post stab and continuing through 24 weeks, with no evidence of spontaneous recovery or reversal of magnetic resonance imaging changes. Radiograph findings included early osteophyte formation by 6 weeks post stab and extensive, bridging osteophytes by 24 weeks. Histologic analysis revealed progressive loss of notochordal cells from the nucleus pulposus, filling of the nucleus pulposus space with fibrocartilage, and derangement of anulus fibrosus. Conclusions. Stabbing the anterolateral anulus fibrosus of adult rabbit lumbar discs with a 16-gauge hypodermic needle to a limited (5-mm) depth results in a number of slowly progressive and reproducible magnetic resonance imaging, radiograph, and histologic changes over 24 weeks that show a similarity to changes seen in human intervertebral disc degeneration. This model would appear suitable for studying pathogenesis and pathophysiology of intervertebral disc degeneration and testing safety and efficacy of novel treatments of intervertebral disc degeneration.

The Natural History of the Anterior Cruciate Ligament-Deficient Knee Changes in Synovial Fluid Cytokine and Keratan Sulfate Concentrations

Restoring knee stability through reconstruction, while providing symptomatic relief, has not been shown to decrease the incidence of degenerative changes after rupture of the anterior cruciate ligament. This suggests that posttraumatic osteoarthritis may not be purely bio mechanical in origin, but also biochemical. To test this, we measured the levels of seven cytokine modulators of cartilage metabolism in knee joint synovial fluid after anterior cruciate ligament rupture. We also measured keratan sulfate, a product of articular cartilage catab olism. The sample population consisted of patients with uninjured knee joints (N = 10), and patients with acute ( N = 60), subacute (N = 18), and chronic (N = 8) anterior cruciate ligament-deficient knees. Synovial fluid samples were analyzed by enzyme-linked immu nosorbent assays. Normal synovial fluids contained high levels of the interleukin-1 receptor antagonist but low concentrations of other cytokines. Immediately af ter ligament rupture there were large increases in in terleukins 6 and 8, tumor necrosis factor α, and keratan sulfate. Interleukin-1 levels remained low throughout the course. As the injury became subacute and then chronic, interleukin-6, tumor necrosis factor-α, and keratan sulfate levels fell but remained considerably elevated 3 months after injury. Concentrations of inter leukin-1Ra fell dramatically. Granulocyte-macrophage colony-stimulating factor concentrations were normal acutely and subacutely but by 3 months after injury were elevated 10-fold. Our data reveal a persistent and evolving disturbance in cytokine and keratan sulfate profiles within the anterior cruciate ligament-deficient knee, suggesting an important biochemical dimension to the development of osteoarthritis there.

Wound-Healing Risk Factors After Open Reduction and Internal Fixation of Calcaneal Fractures

This retrospective study investigated outcomes of wound healing in a series of 63 consecutive patients with 64 fractures of the calcaneus who underwent open reduction and internal fixation done by two surgeons experienced in this fracture during a 3-year period. Thirty-nine patients were managed preoperatively as outpatient referrals before surgery. Twenty-four patients were admitted directly to the trauma service and were managed as inpatients preoperatively. Minimum patient follow-up was 6 months, with an average follow-up of 18 months. A trend correlating the time between injury and operative intervention with the incidence of complications in wounds was noted; the incidence rose in patients who underwent surgery >5 days after their injury. Two-layered closures had a lower incidence of dehiscence compared to single-layered tension-relieving sutures. Patients with a higher body-mass index (BMI) (kg/ m2) took longer to heal their wounds. Strong trends were noted to link BMI and severity of fractures. In the outpatient group, a history of active smoking preoperatively correlated with increased time to wound healing. In 43 patients, there were no wound-healing complications. In 21 feet, there were varying degrees of wound dehiscence. Average wound healing took 47 days. Risk factors for complications in the wound after calcaneal open reduction and internal fixation include single layered closure, high BMI, extended time between injury and surgery, and smoking. Age, type of immobilization, medical illness (including diabetes), type of bone graft, or use of a Hemovac did not influence wound healing.

Bone mineral density and blood flow to the lower extremities: the study of osteoporotic fractures.

This study tests the hypothesis that reduced blood flow to the lower extremities may affect bone remodeling, resulting in a decrease in bone mineral density (BMD). BMD was measured in the axial and appendicular skeleton of 1292 elderly women (mean age, 71 years) enrolled in the Study of Osteoporotic Fractures. The ratio of the posterior tibial and brachial systolic blood pressures, the ankle/arm index, was used as a measure of blood flow to the legs. In the cross‐sectional analysis, this index was positively correlated with BMD at the radius, calcaneus, and hip, but not at the spine. A decrease in the index of 2 standard deviations (SD) (as might occur in patients with moderate arterial disease) was associated with a decrease of 3.7% (95% CI, 1.7%, 5.8%) in hip BMD. The effect size at the hip decreased from 3.7 to 1.8% (and was not statistically significant) when adjustment was made for smoking and/or body mass index (BMI). In the prospective analysis, the rate of bone loss at the hip and calcaneus was greater (p < 0.05) among women whose annual decrease in ankle/arm index was more than 1 SD greater than the mean decrease. This increase was independent of estrogen use, smoking, BMI, pattern of fat distribution, history of diabetes, exercise, and ability to walk. The results from this prospective community‐based study provide the first evidence that among relatively healthy older women decreased vascular flow in the lower extremities may be associated with an increased rate of bone loss at the hip and calcaneus.

Foot and ankle fractures in elderly white women. Incidence and risk factors.

Background: Although foot and ankle fractures are among the most common nonspinal fractures occurring in older women, little is known about their epidemiology. This study was designed to determine the incidence of and risk factors for foot and ankle fractures in a cohort of 9704 elderly, nonblack women enrolled in the multicenter Study of Osteoporotic Fractures. Methods: At their first clinic visit, between 1986 and 1988, the women provided information regarding lifestyle, subjective health, and function. Bone mineral density was measured in the proximal and distal parts of the radius and in the calcaneus. The women were followed for a mean of 10.2 years, during which time 301 of them had a foot fracture and 291 had an ankle fracture. The fractures were classified with use of a modification of the Orthopaedic Trauma Associations guidelines. Results: The incidence of foot fractures was 3.1 per 1000 woman-years, and the incidence of ankle fractures was 3.0 per 1000 woman-years. The most common ankle fracture was an isolated fibular fracture (prevalence, 57.6%), and the most common foot fracture was a fracture of the fifth metatarsal (56.9%). Women who sustained an ankle fracture had been slightly younger at the time of study enrollment than the women who did not sustain such a fracture (71.0 compared with 71.7 years), they had a higher body mass index (27.6 compared with 26.5), and they were more likely to have fallen within the twelve months prior to filling out the original questionnaire (38.1% compared with 29.7%). The appendicular bone mineral density was not significantly different between these two groups of subjects. Women who sustained a foot fracture had a lower bone mineral density in the distal part of the radius (0.345 g/cm 2 compared with 0.363 g/cm 2 ) and a lower calcaneal bone mineral density (0.394 g/cm 2 compared with 0.404 g/cm 2 ) than the women without a foot fracture, they were less likely to be physically active (62.3% compared with 67.8%), and they were more likely to have had a previous fracture after the age of fifty (45.5% compared with 36.8%) and to be using either long or short-acting benzodiazepines. Conclusions: Overall, foot fractures appeared to be typical osteoporotic fractures, whereas ankle fractures occurred in younger women with a relatively high body mass index. Level of Evidence: Prognostic study, Level I-1 (prospective study). See Instructions to Authors for a complete description of levels of evidence.

The Relationship between Ankle‐Arm Index and Mortality in Older Men and Women

To determine whether the ankle‐arm blood pressure index is a useful predictor of mortality in a large group of patients aged 50 or older.

Distal radius fractures in older women: a 10-year follow-up study of descriptive characteristics and risk factors. The study of osteoporotic fractures.

OBJECTIVES: To determine the incidence of distal radius fractures and the characteristics of those fractures and to identify the key risk factors.

Bone Mineral Density and Aortic Calcification: The Study of Osteoporotic Fractures

OBJECTIVE: To investigate the relationship between bone mineral density (in the axial and appendicular skeleton) and calcification of the aorta.