The Change of Bone Metabolism in Ovariectomized Rats : Analyses of MicroCT Scan and Biochemical Markers of Bone Turnover

OBJECTIVE: The purpose of this study was to verify the appropriateness of ovariectomized rats as the osteoporosis animal model. METHODS: Twelve female Sprague-Dawley rats underwent a sham operation (the sham group) or bilateral ovariectomy [the ovariectomy (OVX) group]. Eight weeks after operations, serum biochemical markers of bone turnover were analyzed; osteocalcin and alkaline phosphatase, which are sensitive biochemical markers of bone formation, and C-terminal telopeptide fragment of type I collagen C-terminus (CTX), which is a sensitive biochemical marker of bone resorption. Bone histomorphometric parameters and microarchitectural properties of 4th lumbar vertebrae were determined by micro-computed tomographic (CT) scan. RESULTS: The OVX group showed on average 75.4% higher osteocalcin and 72.5% higher CTX levels than the sham group, indicating increased bone turnover. Micro-CT analysis showed significantly lower bone mineral density (BMD) (p=0.005) and cortical BMD (p=0.021) in the OVX group. Furthermore, the OVX group was found to have a significantly lower trabecular bone volume fraction (p=0.002). CONCLUSION: Our results showed that bone turnover was significantly increased and bone mass was significantly decreased 8 weeks after ovariectomy in rats. Thus, we propose that the ovariectomized rat model be considered a reproducible and reliable model of osteoporosis.

Histomorphometric Analysis of the Spine and Femur in Ovariectomized Rats Using Micro-Computed Tomographic Scan

OBJECTIVE: The purpose of this study was to evaluate the different patterns of bone loss between the lumbar spine and the femur after ovariectomy in rats. METHODS: Twenty-four female Sprague-Dawley rats underwent a sham operation (the sham group) or bilateral ovariectomy (the ovariectomized group). Four and eight weeks after operation, six rats from each of the two groups were euthanized. Serum biochemical markers of bone turnover including osteocalcin and alkaline phosphatase (ALP), which are sensitive biochemical markers of bone formation, and the telopeptide fragment of type I collagen C-terminus (CTX), which is a sensitive biochemical marker of bone resorption, were analyzed. Bone histomorphometric parameters of the 4th lumbar vertebrae and femur were determined by micro-computed tomography. RESULTS: Ovariectomized rats were found to have higher osteocalcin, ALP and CTX levels than sham controls. Additionally, 8 weeks after ovariectomy in the OVX group, serum levels of osteocalcin, ALP and CTX were significantly higher than those of 4 weeks after ovariectomy. Bone loss after ovariectomy was more extensive in the 4th lumbar spine compared to the femur. Bone loss in the 4th lumbar spine was mainly caused by trabecular thinning, but in the femur, it was mainly caused by trabecular elimination. CONCLUSION: The present study demonstrates different patterns of bone loss between the 4th lumbar spine and the femur in ovariectomized rats. Therefore, when considering animal models of osteoporosis, it is important that bone sites should be taken into account.

The Neuroprotective Effect of Treatment of Valproic Acid in Acute Spinal Cord Injury

OBJECTIVE: Valproic acid (VPA), as known as histone deacetylase inhibitor, has neuroprotective effects. This study investigated the histological changes and functional recovery from spinal cord injury (SCI) associated with VPA treatment in a rat model. METHODS: Locomotor function was assessed according to the Basso-Beattie-Bresnahan scale for 2 weeks in rats after receiving twice daily intraperitoneal injections of 200 mg/kg VPA or the equivalent volume of normal saline for 7 days following SCI. The injured spinal cord was then examined histologically, including quantification of cavitation. RESULTS: Basso-Beattie-Bresnahan scale scores in rats receiving VPA were significantly higher than in the saline group (p<0.05). The cavity volume in the VPA group was significantly reduced compared with the control (saline-injected) group (p<0.05). The level of histone acetylation recovered in the VPA group, while it was significantly decreased in the control rats (p<0.05). The macrophage level was significantly decreased in the VPA group (p<0.05). CONCLUSION: VPA influences the restoration of hyperacetylation and reduction of the inflammatory reaction resulting from SCI, and is effective for histology and motor function recovery.

The Neuroprotective Effect of Treatment of Valproic Acid in Acute Spinal Cord Injury

OBJECTIVE: Valproic acid (VPA), as known as histone deacetylase inhibitor, has neuroprotective effects. This study investigated the histological changes and functional recovery from spinal cord injury (SCI) associated with VPA treatment in a rat model. METHODS: Locomotor function was assessed according to the Basso-Beattie-Bresnahan scale for 2 weeks in rats after receiving twice daily intraperitoneal injections of 200 mg/kg VPA or the equivalent volume of normal saline for 7 days following SCI. The injured spinal cord was then examined histologically, including quantification of cavitation. RESULTS: Basso-Beattie-Bresnahan scale scores in rats receiving VPA were significantly higher than in the saline group (p<0.05). The cavity volume in the VPA group was significantly reduced compared with the control (saline-injected) group (p<0.05). The level of histone acetylation recovered in the VPA group, while it was significantly decreased in the control rats (p<0.05). The macrophage level was significantly decreased in the VPA group (p<0.05). CONCLUSION: VPA influences the restoration of hyperacetylation and reduction of the inflammatory reaction resulting from SCI, and is effective for histology and motor function recovery.

Effect of Dietary Calcium on Spinal Bone Fusion in an Ovariectomized Rat Model

OBJECTIVE: To evaluate the effect of calcium supplementation on spinal bone fusion in ovariectomized (OVX) rats. METHODS: Sixteen female Sprague Dawley rats underwent bilateral ovariectomy at 12 weeks of age to induce osteoporosis and were randomly assigned to two groups : control group (n=8) and calcium-supplemented group (OVX-Ca, n=8). Autologous spinal bone fusion surgery was performed on both groups 8 weeks later. After fusion surgery, the OVX-Ca group was supplemented with calcium in drinking water for 8 weeks. Blood was obtained 4 and 8 weeks after fusion surgery. Eight weeks after fusion surgery, the rats were euthanized and the L4-5 spine removed. Bone fusion status and fusion volume were evaluated by manual palpation and three-dimensional computed tomography. RESULTS: The mean fusion volume in the L4-5 spine was significantly greater in the OVX-Ca group (71.80+/-8.06 mm3) than in controls (35.34+/-8.24 mm3) (p<0.01). The level of osteocalcin, a bone formation marker, was higher in OVX-Ca rats than in controls 4 weeks (610.08+/-10.41 vs. 551.61+/-12.34 ng/mL) and 8 weeks (552.05+/-19.67 vs. 502.98+/-22.76 ng/mL) after fusion surgery (p<0.05). The level of C-terminal telopeptide fragment of type I collagen, a bone resorption marker, was significantly lower in OVX-Ca rats than in controls 4 weeks (77.07+/-12.57 vs. 101.75+/-7.20 ng/mL) and 8 weeks (69.58+/-2.45 vs. 77.15+/-4.10 ng/mL) after fusion surgery (p<0.05). A mechanical strength test showed that the L4-5 vertebrae in the OVX-Ca group withstood a 50% higher maximal load compared with the controls (p<0.01). CONCLUSION: Dietary calcium given to OVX rats after lumbar fusion surgery improved fusion volume and mechanical strength in an ovariectomized rat model.

Neurologic and Histological Study of Clip Compression Model for Rat Thoracic Spinal Cord Injuries

OBJECTIVE: The purpose of this study was to assess the time-response of clip compression model for the relationship between the duration of the injury on the rat thoracic spinal cord, and histological and functional outcome measures. METHODS: After laminectomy at T9 in Sprague-Dawley rats, a modified aneurysm clip with a closing force of 30-gram was applied extradurally around the spinal cord at T9, and then rapidly released with cord compression persisting for 1, 5, and 10 minutes, respectively. The locomotor function, according to the Basso-Beattie-Bresnahan (BBB) scale, was assessed weekly for 4 weeks after the injury. The injured spinal cord was then examined histologically including quantification of cavitation. RESULTS: Spinal cord injury by clip compression resulted in worsened BBB scale scores. However, there was spontaneous functional improvement in times for all 3 injury severities, with the greatest improvement in the 1-minute compression group. From 1 week after the injury, BBB scores in the 1-minute group were significantly higher than in the 5 or 10-minutes groups until the end of the follow-up period (p<0.05). For histological analysis, the cavitation area and cavity volume at 4 weeks was directly proportional to the severity of the injury. CONCLUSION: The results of this study show that the rat thoracic cord clip compression model is a reliable and reproducible spinal cord injury model. The duration of clip compression injury in the rat thoracic cord has been correlated with both functional and histologic outcome measures.