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In Vitro Characteristics of Cultured Posterior Longitudinal Ligament Tissue

Epstein, Nancy E., MD,*†; Grande, Daniel A., MD, PhD,†; Breitbart, Arnold S., MD

Cervical Spine
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Study Design.  To determine the osteogenicity of posterior longitudinal ligament ossification, the posterior longitudinal ligament obtained during anterior cervical surgery from patients with the disorder was analyzed with in vitro cultures.

Objectives.  To determine the osteogenicity of the posterior longitudinal ligament.

Summary of Background Data.  The osteogenicity of posterior longitudinal ligament ossification in North America requires better documentation.

Methods.  The posterior longitudinal ligament obtained during anterior cervical corpectomy with fusion from seven patients, three with ossification of the posterior longitudinal ligament documented by magnetic resonance imaging and computed tomography and four with spondylosis, was blindly submitted for in vitro culture. Explants of the posterior longitudinal ligament were placed in Dulbecco modified Eagle medium with 10% fetal calf serum, antibiotics, 4 mmol/L L-proline, and 50 mg/L ascorbic acid. After reaching confluency, cells were trypsinized, and first-passage cells were used for all osteocalcin measurements to establish their osteoblastic phenotype. Periosteal cells, previously shown to synthesize osteocalcin, were used as a positive control. The cells were incubated with 1,25(OH)2 vitamin D3 at 10E-8 M for 72 hours in serum-free medium. The supernatants were collected and frozen, after which the quantity of osteocalcin induced by exposure to 1,25(OH)2 vitamin D3 was determined using enzyme-linked immunoassay. Control replicate cultures were measured without incubation using vitamin D3.

Results.  Ossification of the posterior longitudinal ligament cell lines responded positively with osteocalcin synthesis in the 0.1 to 0.4 ng/M range. The cell line of the patient with spondylosis alone did not respond to vitamin D3 priming.

Conclusions.  Posterior longitudinal ligament cells from the three North American white patients with ossification of the posterior longitudinal ligament, when cultured in vitro, synthesized osteocalcin on vitamin D3 priming, confirming their osteoblastic phenotype, whereas posterior longitudinal ligament cells from four white patients with isolated spondylosis did not.

From the *Albert Einstein College of Medicine, Bronx, New York,

the †North Shore-Long Island Jewish Health System, Manhasset, New York, and New Hyde Park, New York, and

the ‡Division of Plastic and Reconstructive Surgery, Columbia University College of Physicians and Surgeons, New York, New York.

Acknowledgment date: July 21, 2000.

First revision date: April 12, 2001.

Acceptance date: June 4, 2001.

Device status category: 1.

Conflict of interest category: 12,14.

Address reprint requests to

Nancy E. Epstein, MD

Long Island Neurosurgical Associates

410 Lakeville Road, Suite 204

New Hyde Park, NY 11042

E-mail: Don@psych.columbia.edu

© 2002 Lippincott Williams & Wilkins, Inc.