Effect of alamethicin, gramicidin S and melittin upon the particulate guanylate cyclase from rat lung.

The channel-forming antibiotic alamethicin activated rat lung particulate guanylate cyclase (GTP pyrophosphate-lyase (cyclizing) EC 4.6.1.2), and the activated enzyme was further stimulated by sodium nitroprusside when a thiol such as 2-mercaptoethanol was present. Similar effects were seen with the antibiotic gramicidin S and with melittin, a polypeptide purified from bee venom. All of these agents are amphiphilic polypeptides. Nitroprusside was not able to stimulate both particulate and soluble enzyme treated with the nonionic amphiphile, Lubrol PX, suggesting that the membrane-active polypeptides had a different mechanism of action. These polypeptides are known to alter the membrane matrix by binding to phospholipid, and we suggest that this alteration allowed greater access of substrate and of nitroprusside to the enzyme. Lubrol PX, however, may interact preferentially with the enzyme, and thus block nitroprusside activation. The most potent of these agents was melittin, which stimulated nitroprusside activation at a concentration which had little effect by itself (7 microns), and at which others have demonstrated lytic effects on cells.

Regulation of intestinal mucosa guanylate cyclase by hemin, heme and protoporphyrin IX.

Mg2+-dependent activity of intestinal brush border guanylate cyclase was stimulated 4-5-fold by 50-100 microM hemin. Higher concentrations were inhibitory. In the presence of 25% dimethyl sulfoxide, which stimulated activity 9-times, 50 microM hemin further increased activity 1.7-fold. However, when activity was stimulated 32-fold by the Escherichia coli heat-stable enterotoxin, or 26-fold by Lubrol PX, hemin produced only concentration-dependent inhibition. The first type of activation was more sensitive to hemin than the second. Reduction of hemin by dithiothreitol eliminated stimulation of basal activity, while inhibition of Lubrol PX-stimulated activity remained. Protoporphyrin IX also had no effect on basal activity, however, it inhibited enterotoxin- and Lubrol PX-stimulated activities similarly, but only to half the extent of hemin. Substitution of Mn2+ for Mg2+ elevated basal activity 15-fold, and this Mn2+-dependent activity was inhibited by hemin. Mn2+-dependent activity was stimulated (43%) by enterotoxin, however, the stimulated activity was more sensitive to hemin inhibition than the basal Mn2+-dependent activity and both inhibition curves were congruent above 50 microM hemin. Hemin inhibition of Lubrol PX-stimulated activity was much less with Mn2+ than with Mg2+. These results were interpreted as suggesting two sites of hemin inhibition; on an inhibitory regulator and on the enzyme. We also found that the secretory effect of enterotoxin in the suckling mouse bioassay was reduced 56% by the oral administration of hemin.

Kinetic effects of the concentration-dependent stimulation of soluble guanylate cyclase from rat lung.

Soluble guanylate cyclase (GTP pyrophosphate-lyase (cyclizing), EC 4.6.1.2) from rat lung demonstrated concentration-dependent stimulation, that is, an increase in specific activity with increasing enzyme (protein) concentration. This phenomenon persisted through several steps of enzyme purification and was apparently due to the presence of a macromolecular activator, similar in size to the enzyme. Treatment of partially purified enzyme with N-ethylmaleimide destroyed catalytic activity, but did not effect the ability of the preparation to stimulate activity. Kinetic analysis demonstrated that the stimulation was due to an increased V value with no change in the apparent Km value for MnGTP. Stimulation occurred without a time lag, the activator apparently interacting reversibly with the enzyme to increase catalytic capability. Some nonionic detergents of the Triton series inhibited enzyme activity by decreasing the V value, with no change in the Km value, and also decreased concentration-dependent stimulation. However, the two phenomena were not directly related. While the physiological significance of the activator is unclear, its presence affects estimations of recovery during enzyme purification, V determinations, and determinations of the effect of hormone or drug treatment on the activity of tissue extracts.

In vitro hyperextension injuries in the human cadaveric cervical spine.

To investigate the relationship between the type of hyperextension injuries and the conditions producing them, nine cervical specimens (occiput to T1) were loaded to failure in tension at a fixed extension angle of 30 degrees. Under these loading conditions, specimens failed at average tensile loads and extension moments of 499 +/- 148 (SD) N and 4.0 +/- 3.1 Nm, respectively. Failure occurred at an average tensile displacement of 18.8 +/- 7.7 mm. The anterior longitudinal ligament ruptured and the intervertebral disc failed in at least one level in all specimens. In four specimens, the disc failed at an additional level, leaving the anterior longitudinal ligament intact at that site. With one exception, all injuries occurred in the lower cervical spine (C5-C6 and C6-C7), the region most often injured in vivo. The location of the injuries was associated with the degree of degeneration of the facet joints and the discs. The discs of the lower cervical spine were significantly more degenerated than those at the C2-C3 level. In addition, the degree of disc degeneration in the noninjured discs was significantly less than in the injured discs. These data help quantify the threshold of injury and the patterns of tissue damage resulting from hypertension loading of the cervical spine.

Monitoring the healing of a tibial osteotomy in the rabbit treated with external fixation.

Fracture site axial rigidity was monitored non-destructively at weekly intervals during healing of tibial osteotomies in adult rabbits. Two groups of 20 rabbits each were treated with external fixators of two different rigidities. Four animals from each group were killed at 3, 5, 6, 7, and 8 weeks to determine the bending moments at failure of the healing fractures. Normal fracture healing was accompanied by characteristic phases in the development of fracture site axial rigidity. From 0 to 3 weeks there was a period of low and approximately constant rigidity, followed by a linear increase during 3 to 5 weeks to an approximately three to four times greater rigidity. The maximum average normalized axial rigidities were reached at 6 weeks and were 57% (high rigidity group) and 77% (low rigidity group) of the untreated contralateral controls. The maximum average normalized failure moments occurred at 8 weeks and were 48% (high rigidity) and 44% (low rigidity) of controls. The differences due to fixator rigidity were not statistically significant except for a large increase in failure moments at 3 weeks for the low rigidity group. Axial rigidities were correlated (r2 = 0.74 and 0.53, respectively) with failure moments, but only during the first 6 weeks. The monitoring technique provides a nondestructive means for following the biomechanical progress of fracture healing in an animal model. The occurrence of the characteristic increase in fracture site axial rigidity at 3 to 5 weeks can also be used to distinguish between normal and abnormal healing.

Correlations of radiographic analysis of healing fractures with strength: a statistical analysis of experimental osteotomies.

The assessment of fracture healing strength using routine roentgenograms is difficult and controversial. There are few experimental data that correlate radiographic appearance with the actual quantitative strength of healing fractures. However, this method is widely used in clinical practice. A study is presented in which transversely osteotomized rabbit tibiae were allowed to heal for 3 to 8 weeks. A pair of orthogonal roentgenograms was taken of each bone and the bones were tested for strength in a dynamic torsion testing machine. Statistical analyses were done to study the correlations between the roentgenographic and strength parameters. Cortical continuity was found to be the best single predictor of strength of a healing fracture (correlation coefficient r = 0.80). The least important predictor was the callus area (r = 0.17). Fracture displacement, callus thickness, and callus diameter had negative correlations. From these experimental findings in an animal model, we conclude that even under laboratory conditions the information gained from plain radiographs is not sufficient to accurately predict the strength of a healing fracture.

An epidemiologic study of lifting and twisting on the job and risk for acute prolapsed lumbar intervertebral disc.

An epidemiologic case-control study undertaken in Connecticut during 1979-1981 indicated that persons with jobs requiring lifting objects of more than 11.3 kg (25 lb) an average of more than 25 times per day had over three times the risk for acute prolapsed lumbar intervertebral disc as people whose jobs did not involve lifting objects of this weight. If the body was usually twisted while the lifting was done, this elevation in risk was apparent with less frequent lifting. An especially high risk for prolapsed lumbar disc was associated with jobs involving lifting objects of more than 11.3 kg with the body usually twisted and the knees not bent while the lifting was done. Neither lifting objects of less than 11.3 kg nor twisting without lifting was associated with an increase in risk.

Basic biomechanics of the spine.

The purpose of this presentation is to provide basic biomechanical information concerning the spine, its components, and the spinal cord. It is shown that this information is helpful in understanding the fundamental functions of the spinal column. The experimentally determined physical properties of the vertebra, various spinal ligaments, the disc, and the spinal cord under many different loading conditions are described. The role of the special characteristics of the spine ligaments in allowing physiological motions of the spine, preventing excessive motions between vertebrae, and protecting the spinal cord during trauma are discussed. Movements of the spinal cord within the spinal canal and associated changes of its section during physiological movements of the spine are also described. The kinematics of the various regions of the spine are discussed and their clinical significance is presented. The problems of spinal trauma and is associated spinal instability are analyzed. Guidelines are recommended to assess spinal stability. The proper application of such guidelines will provide the basis for sound clinical judgments.

Fractures of the dens. A multicenter study.

The treatment of fractures of the dens is often inadequate, and surgeons are divided in their opinions regarding the best surgical management of these potentially serious injuries. Because of these concerns, the Cervical Spine Research Society conducted a multicenter survey of its membership regarding the management of these fractures. Fractures of the dens can be effectively classified according to the anatomical level of the fracture, as described by Anderson and d'Alonzo. We have found that the degree of angulation and amount of displacement are also important factors. Fractures occurring at the junction of the dens with the vertebral body (Type-II fractures) were found to be the most troublesome. The initial management of these fractures with a halo device was successful in only 68 per cent; however, posterior cervical fusion was successful in 96 per cent, and that appears to be the treatment of choice. Fractures extending into the vertebral body (Type-III injuries) were found not to be as benign as has been reported. Malunion and non-union occurred in patients with this injury who were treated with an orthosis alone, and a halo device or surgery may be indicated for unstable lesions.

The impact of musculoskeletal disorders on the population of the United States.

Musculoskeletal disorders constitute such a large component of impairments of individuals in the working and geriatric population that good statistical data on them are essential for planning health care. We have collected the pertinent data, with references to recent source material, to determine the incidence, effect on the quality of life, and associated costs of these disorders in the United States.

The four biomechanical stages of fracture repair.

Based on analysis of the torque-angle curves and roentgenographic findings in fifty-three healing tibial fractures in rabbits tested in torsion to failure, four biomechanical stages of fracture healing were defined, as follows: Stage I--failure through original fracture site, with low stiffness; Stage II--failure through original fracture site, with high stiffness; Stage III--failure partially through original fracture site and partially through intact bone, with high stiffness; and Stage IV--failure entirely through intact bone, with high stiffness. These stages correlated with the progressive increases in the average torque and energy absorption to failure as healing progressed and also with the average times since the original experimental fracture. It is hoped that this system of staging will provide both a standard by which important variables related to ultimate strength of healing fractures can be correlated and an objective way to predict delayed unions and non-unions and to determine the level of activity that is safe for patients with a healing fracture.

Mechanical properties of the human thoracic spine as shown by three-dimensional load-displacement curves.

The mechanical properties of the human spine are best described by load-displacement curves which include coupling effects. Three-dimensional load-displacement curves were obtained for all levels of the human thoracic spine using fresh cadaver spines in an atmosphere containing 100 per cent humidity at 22 degrees centigrade to stimulate a physiological environment. Six forces and six moments were applied, one at a time, to the center of the upper vertebra while its subadjacent fellow was fixed. Assuming sagittal plane symmetry, vertebral displacement was measured in three-dimensional space and load-displacement curves were plotted for the main as well as the coupled motions. The thirty-six curves necessary to define the mechanical characteristics of each motion segment completely were determined for all eleven thoracic levels. The curves showed that all the thoracic spine is a complex three-dimensional structure with coupled motion characteristics. Axial forces (compression/tension) resulted in significant horizontal displacements. Spine motion segments were more flexible in flexion than in extension. The spine was found to be least flexible during axial compression.

Comparison of cyclic loading versus constant compression in the treatment of long-bone fractures in rabbits.

UNLABELLED: The strengths of healing long-bone fractures treated in two different mechanical environments were compared using a rabbit experimental model. Constant compression was applied to one healing tibial fracture, while the other was subjected to cyclic compression. At six weeks of healing, the group of tibial fractures treated with cyclic loading exhibited significantly higher torque and energy absorption to failure and lower stiffness than their pair-mates treated with constant compression. No statistically significant differences were detected at four or eight weeks of healing, although there was a suggestion that compression-treated bones may be stronger in the earlier phases of healing. CLINICAL RELEVANCE: This experiment suggests that the requirements for rapid fracture-healing may vary with the stage of healing; that is, rigid immobilization applied during the initial stages of healing followed by intermittent compression during later stages.

Prediction of vertebral body compressive fracture using quantitative computed tomography.

We performed quantitative computed tomography in vitro on the first and third lumbar vertebrae in human cadavera using a dibasic potassium phosphate phantom for calibration. The quantitative computed-tomography numbers exhibited a significant positive correlation (R2 = 0.89, p less than 0.0001) with direct measurements of the apparent density of the vertebral trabecular bone. We also conducted uniaxial compression tests to failure of the vertebral bodies after removal of the posterior elements, and found that vertebral compressive strength was also correlated at a high level of significance (R2 = 0.82, p less than 0.0001) with direct measurement of the trabecular apparent density. These findings suggested the possibility that the quantitative computed-tomography values might be directly predictive of vertebral compressive strength. However, when we correlated the quantitative computed-tomography values directly with vertebral compressive strength, the results (R2 = 0.46, p less than 0.061) were suggestive but not quite significant. All vertebral bodies failed by compression of the end-plate, suggesting only a modest structural role for the cortical shell under these loading conditions. This was confirmed by comparing the compressive load to failure of twenty additional pairs of vertebrae that were tested with and without an intact vertebral cortex. Removal of the cortex was associated with approximately 10 per cent reduction in vertebral load to failure.

An epidemiological study of acute prolapsed cervical intervertebral disc.

In this epidemiological study of acute prolapsed cervical intervertebral disc, we found that people in the fourth decade of life were affected somewhat more frequently than individuals in other age groups, and men with a prolapsed cervical disc outnumbered women by a ratio of 1.4 to one. Factors that were associated relatively strongly with this diagnosis were frequent lifting of heavy objects on the job that was held around the time of the onset of symptoms, cigarette-smoking, and frequent diving from a board. Positive associations that were of borderline statistical significance or were not statistically significant were found with operating or driving vibrating equipment and time spent in motor vehicles. Variables that did not appear to affect the risk for a prolapsed cervical disc included participation in certain sports other than diving, frequent wearing of shoes with high heels, the number of pregnancies or live births, frequent twisting of the neck on the job, time spent sitting on the job, and smoking cigars or a pipe.

Variations of stiffness and strength along the human cervical spine.

The load-displacement response and strength of the mid (C2-C5) and lower (C5-T1) cervical regions were determined for combinations of sagittal loads, in vitro. In unpaired t-test comparisons, the mid cervical region was significantly stiffer in compression and extension than the lower region. In tests to failure, failure in six out of seven mid cervical specimens resulted from flexion alone, while combined compression-flexion was required to fail five of the eight lower cervical specimens. Post-test dissections revealed no regional differences in the pattern of failure. In addition to sagittal tests, the load-displacement responses of three-vertebrae cervical specimens were measured with the upper body axially rotated with respect to the lower body. The effect of this pre-torsion was to diminish the zone of low slope near zero load for axial, shear, and flexion motion. Three of the four axially rotated specimens failed in flexion without added compression. These controlled load-displacement measurements of cervical spine specimens describe for the first time the continuous flexion-compression response up to failure, and suggest that consideration of the biomechanics of three apparently distinct mobile regions of the cervical spine (C1-C2, C2-C5, C5-T1) may facilitate the interpretation of hazardous conditions and the diagnosis of injury. These data also provide basic information for the in vitro investigation of passive cervical spine protection such as helmets and head-rests, suggesting that the head should be kept in a non-rotated position to reduce risk of injury.

Physical and mechanical properties of calf lumbosacral trabecular bone.

The physical and mechanical properties of calf lumbar and sacral trabecular bone were determined and compared with those of human trabecular bone. The mean tissue density (1.66 +/- 0.12 g cm-3), equivalent mineral density (169 +/- 36 mg cm-3), apparent density (453 +/- 89 mg cm-3), ash density (194 +/- 59 mg cm-3), ash content (0.6 +/- 0.05%), compressive strength (7.1 +/- 3.0 MPa) and compressive modulus (173 +/- 97 MPa) of calf trabecular bone are similar to those of young human. There were moderate, positive linear correlations between apparent density and equivalent mineral density, ash density, and compressive strength; and between compressive strength and equivalent mineral density (R2 ranging from 0.35 to 0.48, p less than 0.001). Apparent density, ash density, and equivalent mineral density did not differ significantly in different regions. In contrast to humans, the compressive strength increased from posterior, near the facet, to the anterior vertebral body. These comparisons of physical and mechanical properties, as well as anatomical comparisons by others, indicate that the calf spine is a good model of the young non-osteoporotic human spine and thus useful for the testing of spinal instrumentation.

Stress analysis of compression plate fixation and its effects on long bone remodeling.

A three-dimensional finite element model is generated for an intact plexiglass tube with an attached six-hole stainless steel compression plate. The results for a wide range of loads, including cyclic external loads and static tensile preloads in the plate and screws, are examined as specifically related to plate-induced osteopenia. The model demonstrates that disuse osteopenia, resulting from a reduction in magnitude of cyclic axial stress, should be limited to the central region between the inner screws. Also, the addition of a static preload negates any reduced axial stress levels in this region, thus raising questions on the relative importance of static and cyclic stresses for the internal remodeling of bone.

Biomechanical considerations in the surgical management of cervical spondylotic myelopathy.

Based on a review of previous studies and our opinion, biomechanical considerations suggest the following guidelines for the surgical management of CSM. It is not recommended that the dura mater, the pia mater, and the dentate ligaments be transected in the surgical treatment of CSM. Anterior decompression and fusion, preferably with the Smith-Robinson technique, is recommended for patients with anterior impingement of the spinal cord at one or two levels in the absence of a narrow spinal canal. This procedure is also advantageous when there is significant radiculopathy associated with the level(s) of pathology. Posterior decompression is recommended when there are three or more levels involved, and particularly when there is developmental stenosis of the canal, ie, a DAD below 13 mm and a SAD below 11 mm. Laminectomy and laminoplasty for CSM may not be any different as regards surgical outcome. One well-controlled study showed only one difference, a decrease in the ability of the laminoplasty patients to extend the neck. If there is evidence of instability or a potential for it, posterior decompression procedures should be accompanied by a facet fusion, or in the case of laminoplasty, some fusion modification such as that described by Itoh and Tsuji. There may also be circumstances in which significant multilevel anterior spur formation and compression in association with a stenotic canal should be treated with anterior and posterior surgery with appropriate attention to maintaining adequate stability. The advantages and disadvantages of these various surgical procedures and their relative appropriateness in various clinical situations will be gradually clarified through well-designed and executed laboratory and clinical investigations.

Epidemiology and impact of low-back pain.

Disorders of the lumbar spine are among the most common medical problems in western countries, affecting up to 80% of people at some time during their lives. The epidemiology and impact of six specific disorders of the lumbar spine are reviewed. These include prolapsed discs, disc degeneration, osteoarthrosis of the apophyseal joints, fractures and dislocations of vertebrae, osteoporosis, and spondylolisthesis. Various mechanical factors contribute to the causation of most of these disorders, but other underlying pathologic mechanisms are important as well. In light of the great impact of these conditions on society and on individuals, it is concluded that there is a considerable need for a greater allocation of resources for improvement in methods of prevention, diagnosis, and treatment.