Hormone replacement therapy prevents osteoclastic hyperactivity: A histomorphometric study in early postmenopausal women.

In a randomized, double blind, clinical prospective trial comprising 35 women treated with either hormone replacement therapy (HRT) (cyclic estradiol/norethisterone acetate) or placebo we performed histomorphometric studies on paired bone biopsies obtained before and after 2 years of treatment. Untreated women developed a progressively more negative balance at individual bone multicellular units (BMUs) (i.e., wall thickness-erosion depth) (2.2 +/- 1.7 microm vs. -5.7 +/- 1.4 microm; p < 0.01), while women on HRT displayed preservation of bone balance (2.4 +/- 2.4 microm vs. 2.5 +/- 2.5 microm; NS). No significant differences in wall thickness between the two groups were demonstrable, but the untreated women developed a pronounced increase in erosion depth over 2 years (46.9 +/- 1.8 microm vs. 52.0 +/- 1.9 microm; p < 0.05), while the HRT group revealed no change (47.8 +/- 2.7 microm vs. 44.6 +/- 1.7 microm; NS). Furthermore, the placebo group displayed an increased osteoclastic erosion depth (17.8 +/- 1.6 microm vs. 25.0 +/- 1.7 microm; p < 0.001), compared with unchanged values in the HRT group (20.0 +/- 1.6 microm vs. 16.9 +/- 1.4 microm/day; NS). While the placebo group revealed a slight increase in volume referent resorption rate (35 +/- 8% vs. 38 +/- 8%; NS) the HRT group revealed a pronounced decrease (46 +/- 8% vs. 28 +/- 5%; p < 0.05). No significant changes in marrow star volume (an index of trabecular perforations) were demonstrable in either group. Our results demonstrate that bone remodeling in early postmenopausal women is characterized by progressive osteoclastic hyperactivity, which is reduced by cyclic HRT. This reduction of resorptive activity at the BMU level after HRT seems to precede the reduction in activation frequency demonstrated in previous studies on older postmenopausal women.

Star volume of marrow space and trabeculae in iliac crest: sampling procedure and correlation to star volume of first lumbar vertebra.

The star volume is a new stereologic parameter which can describe structural changes of trabecular bone. It is defined as the mean volume of all the parts of an object which can be seen unobscured in all directions from a particular point inside the object. The mean is over all points inside the object. The parameter is only unbiased when Baddeley's vertical or isotropic sections are used. The star volume of the marrow space (V*m.space) and of the trabeculae (V*tr) was estimated on vertical sections obtained from the first lumbar vertebra and the iliac crest. The continued autopsy material presented was from 7 females and 11 males without malignant or metabolic bone diseases. The mean age was 52 years (range 7-87 years). The sampling procedure was as required for vertical sections and an anisotropic test system was used. A significant age-related increase was shown in marrow space star volume in both lumbar vertebra and iliac crest. The increase in lumbar vertebra marrow space star volume was significantly greater for women than for men. Moreover, a significant correlation was found between the marrow space star volume in lumbar vertebra and iliac crest (r = 0.72, 2p less than 0.01). No such relation was found for trabecular star volume which did not change with age. The results also revealed that marrow space star volume could be measured on conventional bone biopsies preferably, however, on two mutually perpendicular vertical section planes.

Star volume of marrow space and trabeculae of the first lumbar vertebra: sampling efficiency and biological variation.

The decrease in the amount of trabecular bone which is seen with age cannot solely be explained by thinning of trabeculae but must also be due to a loss of structural trabecular bone leading to a discontinuity in the trabecular network. Due to the complex architecture and the anisotropy of bone it is difficult to demonstrate this structural change by conventional histomorphometry. Unbiased stereological estimators can however be obtained from anisotropic structures when using vertical sections and a specially designed anisotropic test system. This combined with a new and unbiased stereological parameter for bone structure the star volume can be of major importance in clarifying histological changes of bone structure. The star volume is defined as the mean volume of all the parts of an object which can be seen unobscured in all directions from a particular point with the mean value taken over all points inside the object. It is defined for any type of objects including cavities like marrow space and networks like the trabecular system. Measurements are performed using a frame and a grid with points and lines. The material investigated was the first lumbar vertebra obtained from two females and six males with ages 26 to 75 years without malignant or metabolic bone diseases. The sampling procedure was as required for vertical sections. Results did show a highly significant, five-fold increase in the star volume of the marrow space with age; no such age correlation was found for the star volume of the trabeculae. The only explanation for such an increase in the size of the marrow space is by removing or perforating trabecular bone.(ABSTRACT TRUNCATED AT 250 WORDS)

Unbiased stereological estimation of osteoid and resorption fractional surfaces in trabecular bone using vertical sections: sampling efficiency and biological variation.

The conventional stereological procedure for estimating bone surface densities in iliac crest biopsies provides biased estimates because bone substructure is anisotropic and the sectioning of the biopsies is intended to be parallel to the cylindrical axis of the iliac crest biopsy. It has recently been shown that random anisotropic sections with an identifiable axis or direction which is arbitrary but fixed can be used for unbiased estimates of surface areas. The arbitrary axis is called "vertical," a denomination which does not imply anything about its relation to directions in the world's field of gravity. Sampling efficiency and biological variation of fractional osteoid and resorptive surfaces were investigated in 14 paired bone specimens obtained from two females and five males (mean age 64) using an integral anisotropic test system constructed of cycloids. Results obtained at direct microscopy were compared to results obtained with projection onto the table. Mean fractional osteoid and resorption surfaces were 6.2 and 1.4% using direct microscopy and 5.6 and 1.7% using projection. Estimation of variance at each level of sampling showed that the majority of the total observed variance was due to true variance between individuals; for fractional osteoid surface the unavoidable biological variation was 80.5% of the observed variation and for fractional resorption surfaces it was 57%. The efficiency of the estimation procedure is therefore close to optimal and the only way to improve the precision in groups of patients and controls would be to include more individuals in the study.(ABSTRACT TRUNCATED AT 250 WORDS)

Marrow space star volume in the iliac crest decreases in osteoporotic patients after continuous treatment with fluoride, calcium, and vitamin D2 for five years.

Iliac crest biopsies in osteoporotic patients were obtained before and after five years of continuous treatment with sodium fluoride (40-60 mg/d), vitamin D2 (18,000 IE/d), and calcium phosphate (45 mmol/d). Star volume of marrow space and trabeculae, mean trabecular thickness, and fractional amount of trabecular bone were estimated on so-called vertical sections before and after treatment. Results showed a significant decrease in marrow space star volume after five years of treatment (35 mm3 versus 19 mm3) (2p less than 0.02). No significant changes were observed in trabecular star volume (0.149 mm3 versus 0.104 mm3) (2p greater than 0.50) or mean trabecular thickness (85 microns versus 79 microns, 2p greater than 0.30). A slight but insignificant increase in the fractional amount of trabecular bone was found (9.7% versus 10.4%, 2p greater than 0.50). A rather close inverse correlation was obtained, however, between individual changes in trabecular bone volume fraction and marrow space star volume (r = -0.72, 2p = 0.011). The findings suggest that the large marrow "cavities" had been split into two or more "cavities" by some newly generated trabecular structures leading to increased trabecular connectivity. The strength of trabecular bone may be increased due to this beneficial effect of fluoride on trabecular bone structure.

Unbiased stereologic estimation of surface density in bone using vertical sections.

The conventional stereologic procedure for estimating bone surface densities in iliac crest biopsies gives biased estimates because bone substructure is anisotropic and the sectioning of the biopsies is intended to be parallel to the cylindrical axis of the iliac crest biopsy. It has recently been shown that random anisotropic sections with an identifiable axis or direction that is arbitrary but fixed can be used for unbiased estimates of surface areas. The arbitrary axis is called "vertical," which does not imply anything about its relation to direction of gravity. To describe the methodology and the practical sampling procedure for vertical sections, surface density of trabecular bone was estimated in 16 bone samples obtained at autopsy from eight persons (mean age 64 years) using an anisotropic cycloid test system designed for vertical sections. In addition, variation in the estimation procedure was quantitated. The average surface density of trabecular bone was 2.8 +/- 0.4 mm-1 (+/- SD). No systematic difference was observed between the two bone specimens from each of the eight persons. Estimation of the variance at each level of sampling showed that the majority of the total observed variance was due to true variance between individuals. Vertical section is, in general, the method of choice when dealing with surfaces not known to have an isotropic orientation distribution. The cycloid test system is easy and fast to use, and the estimate is truely unbiased. The requirements are all sections must be parallel to the vertical axis, the rotation around this axis must be random, the position of the section or of the biopsy must be random, and the vertical axis must be identified in the section.

A histomorphometric determination of iliac bone structure and remodeling in obese subjects.

Quantitative histomorphometric analyses of iliac crest biopsies were performed after tetracycline double labeling in 24 patients with morbid obesity and in 30 age- and sex-matched controls. The amount and structure of bone were determined from measurements of total biopsy length, fractional length of medullary space, fractional trabecular bone volume, trabecular thickness, and the intertrabecular distance. Static and dynamic variables of bone resorption and formation were determined, and the balance of the BMU level was estimated from final resorption depth and mean wall thickness of trabecular structural units. In the obese patients the total biopsy length was increased, with a normal proportion of medullary space to total biopsy length. The mean fractional trabecular bone volume was reduced due to an increased distance between trabeculae of normal mean thickness. The total biopsy length in the obese patients was found to be positively related to the intertrabecular distance and inversely related to the fractional trabecular bone volume. The remaining histomorphometric variables describing bone formation rate at tissue, BMU, and cellular levels, the amount of bone formed, the mineralization process, mineralization lag time, bone resorption, and the balance between resorption and formation were all normal in the obese group.

Normal postmenopausal women show iliac crest trabecular thickening on vertical sections.

Trabecular thickness and the trabecular star volume were estimated in 49 normal individuals (20 males and 29 females) using an unbiased sampling procedure consisting of anisotropic, vertical sections, and a corresponding anisotropic test system. Eight-microns-thick undecalcified stained vertical sections were obtained from iliac crest specimens. Implying a trabecular plate model mean trabecular thickness (Tb.Th., microns) was estimated by different methods: (a) the ratio between bone volume and bone surface (Tb.Th.ratio); (b) the mean of the orthogonal intercepts multiplied with pi/4 (Tb.Th.l0); and (c) the mean of random linear intercepts divided by 2 (Tb.Th.l1). Trabecular star volume (Vtr*, mm3), a true three-dimensional size parameter independent on any model, was estimated using linear intercepts in random directions. Significant (2p less than 0.001) correlations were found between the different methods with coefficients of correlation ranging from 0.71 to 0.89. The Tb.Th. ratio was overall higher (155 +/- 31 microns, (X, SD), 2p less than 0.001) than the other linear estimates, Tb.Th.l0: 144 +/- 26 microns and Tb.Th.l1: 135 +/- 27 microns. Estimates based on orthogonal intercepts (l0) were slightly higher (2p less than 0.001) than those from random linear intercepts (l1). Among all the individuals no significant age-related changes were seen in the measured parameters. However, postmenopausal women (greater than 50 years of age) had more thick trabeculae than younger women (2p = 0.03). Intra-individual distribution analysis disclosed that this finding could not be explained by preferential perforation and loss of thinner trabeculae during bone remodeling. Therefore, normal postmenopausal women do increase trabecular thickness with age.(ABSTRACT TRUNCATED AT 250 WORDS)

Biologically meaningful determinants of the in vitro strength of lumbar vertebrae.

Applying unbiased stereological methods and a new stereological parameter, star volume of cancellous bone, the bone structure of the first vertebral body was examined and compared with the compressive strength of the second lumbar vertebra. The material came from eight males, aged 33-69 years (mean 49 years) and seven women, aged 22-87 years (mean 52 years) without malignant or metabolic bone disease. From these individuals, first and second lumbar vertebral body were obtained at autopsy. The heights and weights of the individuals were recorded. The following structural parameters were estimated on undecalcified, seven-microns, Goldner-Trichrome stained vertical sections: fractional volume of trabecular bone (BV/TV%), mean trabecular thickness (Tb.Th.l1 microns), trabecular star volume (V*tr mm3), marrow space star volume (V* m.space mm3), and mean thickness of the lateral cortical ring (microns). The compressive strength of whole vertebral body, mean cross sectional area (cm2), and ash density (g/cm3) were estimated and the data were compared to bone histomorphometric estimates. A significant decrease with age for all parameters was found except for marrow space star volume, which increased. With compressive strength as the dependent variable and all other parameters as independent variables, it was shown by standard multiple regression analysis that the in vitro tested compressive strength could be predicted from mean cortical thickness, mean cross sectional area, and marrow space star volume or ash density with a multiple, squared coefficient of regression (r2) of 0.95 when the height and sex of the individual were known.(ABSTRACT TRUNCATED AT 250 WORDS)

Marked changes in iliac crest bone structure in postmenopausal osteoporotic patients without any signs of disturbed bone remodeling or balance.

Successful iliac crest bone biopsies were obtained from 63 women with postmenopausal vertebral crush fracture osteoporosis. Structural and static histomorphometric parameters were compared with 25 age-matched normal females, who had suffered an unexpected and sudden death. The control group for dynamic parameters comprised 13 younger normal females. Marked structural changes were observed in the osteoporotic patients in cortical as well as cancellous bone. Cortical width, trabecular volume, trabecular bone surface density and trabecular number were all reduced, whereas trabecular separation and star volume were increased. On the other hand trabecular thickness was normal in the patients. These structural changes in cancellous bone indicate that extensive perforations of trabecular plates have occurred or that whole trabecular elements have been removed. The remodeling cycles of cancellous bone surface and the frequency by which they were repeated (activation frequency) did not differ significantly between osteoporotic patients and normal younger women. The bone balance per remodeling cycle in osteoporotic patients and controls was not significantly different. No subset of individuals in the group of osteoporotic patients could be identified regarding extent of resorptive and formative surfaces, bone formation rate or activation frequency. In the present osteoporotic patients nothing in the ongoing remodeling process could explain the marked changes in bone structure. The pathophysiological changes leading to osteoporosis may therefore occur earlier in life, maybe long before the manifestation of the disease.

Primary hyperparathyroidism: iliac crest trabecular bone volume, structure, remodeling, and balance evaluated by histomorphometric methods.

Iliac bone biopsies from 69 patients (48 females, 21 males; median age 58 years; range 17-79 years) with primary hyperparathyroidism (PHP) were examined, and static histomorphometric parameters compared to 30 age- and sex-matched normal controls. The control group for the dynamic parameters constituted 20 sex-matched younger normal controls. Fractional volume of trabecular bone was normal, but the trabeculae were thinner (p less than 0.05) in PHP. The structural parameters marrow space star volume, intertrabecular distance, and mean trabecular plate density were not significantly different in PHP patients compared to normal controls, but the age-related increase, for females, in marrow space star volume and decrease, for both sexes, in mean plate density observed in the controls were not noticed in the PHP group. Trabecular bone remodeling was found significantly increased in the PHP patients reflected by increased extension of eroded (p less than 0.001), osteoid (p less than 0.001), and labeled surfaces (p less than 0.05). The activation frequency was increased by approximately 50% (p less than 0.001). Neither PHP patients nor controls showed age-related decrease in trabecular thickness, and accordingly in both groups the bone balance per remodeling cycle was very close to and not significantly different from zero. Normal postmenopausal women (age greater than or equal to 50 yr) had lower trabecular bone volume (p less than 0.001) and higher intertrabecular distance than normal pre-menopausal women (age less than 50 yr).(ABSTRACT TRUNCATED AT 250 WORDS)

Low bone mineral density in the femoral neck of medieval women: a result of multiparity?

An archaeological investigation of a medieval cemetery gave us the opportunity to investigate 49 Danish skeletons dating from 1000 to 1250 A.D. and to compare them with 298 contemporary Danes (aged 19-79 years) and assess the millennial trend in bone mineral density (BMD) in populations considered genetically closely related. BMD and bone mineral apparent density (BMAD) of the femoral neck were measured by dual-energy X-ray absorptiometry (DEXA) and transformed into z scores. BMD(zscore) was significantly lower in medieval women (-0.54 +/- 0.25, p = 0.04), whereas BMD(zscore) in medieval men was significantly higher (0.55 +/- 0.22, p = 0.02). In medieval women, BMD(zscore) tended to increase with age (r = 0.42, p = 0.07), whereas no change was seen in men (r = 0.19, not significant [n.s.]). Also, BMAD(zscore) was significantly elevated in medieval men (1.00 +/- 0.28, p < 0.01), but in medieval women no difference was found (-0.28 +/- 0.21, n.s.). However, the correlation between BMAD(zscore) and age was significant in the medieval women where it increased with advancing age (r = 0.49, p = 0.03). In conclusion, medieval women had lower BMD when compared with contemporary women, but this relationship was reversed in women who survived to older ages. In contrast, medieval men had significantly higher BMD as compared with contemporary men at all ages. The observed lower BMD in medieval women can be explained by the well-known selective mortality among the younger women. A high birth rate and prolonged periods of lactation are the main reasons for the observed increased mortality, and therefore can also very likely explain the associated low BMD. The increase in the incidence of osteoporosis in modern elderly women could possibly, or partially, be explained by the survival of women who would have died prematurely had they lived in earlier centuries.

Bone histomorphometry in hypoparathyroid patients treated with vitamin D.

Parathyroid hormone (PTH) and vitamin D are both active regulators of bone remodeling. Several studies, mostly in animals and in vitro, have suggested that the two hormones act synergistically or interdependently. The aim of the present study was therefore to describe the actions of vitamin D alone on bone remodeling in the absence of circulating PTH. Bone biopsies were obtained from 12 patients with vitamin D-treated hypoparathyroidism and from 13 age- and gender-matched normal controls. Mean total resorption rate was reduced (0.9 vs. 3.8 mu m/day,p < 0.001), the resorption period was prolonged (80.8 vs. 25.7 days, p < 0.001), and the resorption depth was reduced (41.7 vs. 55.3 mu m, p < 0.001). The fractional active and the total eroded surface were not significantly reduced. The fractional formation surface was reduced (5.2 vs. 12.5 mu m, p < 0.001). Trends toward prolongation of the formation period and reduction of the final wall thickness were found. The balance between resorption depth and final wall thickness was not significantly different from normal (0.96 vs. -4.4 mu m). The quiescent period was prolonged (7.6 vs. 1.7 years, p < 0.001) and the activation frequency was reduced (0.13 vs. 0.6 year(-1), p < 0.001). The structural parameters, trabecular bone volume, trabecular thickness, marrow space star volume, and trabecular star volume, remained unchanged. In the absence of PTH, Vitamin D alone is not able to normalize bone resorption and bone turnover in hypoparathyroid patients.

Primary hyperparathyroidism: bone structure, balance, and remodeling before and 3 years after surgical treatment.

In 19 patients with primary hyperparathyroidism (PHPT) (14 women and 5 men; age 53 +/- 11 years, range 29-69 years), bone densitometry, biochemical markers of bone turnover, and iliac crest bone biopsies were obtained before and 3 years after successful surgical treatment. A significant increase in bone mineral content (BMC) was observed in both the lumbar spine (p < 0.001) and the proximal part of the distal forearm (p < 0.001), whereas the increase in BMC in the femoral neck was insignificant. Biochemical markers of bone formation (serum alkaline phosphatase, serum bone alkaline phosphatase and serum osteocalcin) and resorption (serum pyridinoline cross-linked telopeptide of type I collagen and urine N-telopeptide of type I collagen) all decreased following treatment. In cortical bone, relative cortical width increased following surgery (p < 0.05) and cortical porosity decreased (p < 0.01). No changes were observed in core width or cortical width. In cancellous bone, no significant changes were observed in any of the measured structural parameters. However, significant reductions in the extent of osteoid- (p < 0.01) and tetracycline-labeled surfaces (p < 0.001), and in bone formation rate (p < 0.001) and activation frequency (p < 0.001), were found. The numerical decrease in the extent of eroded surfaces did not reach significance (p = 0.057). No changes were observed in mineral appositional rate and adjusted appositional rate. The amount of bone resorbed (expressed as the resorption depth) and the amount of bone reformed (expressed as wall thickness) per remodeling cycle seemed unaffected by the treatment. Consequently, no effect on bone balance per remodeling cycle could be detected. The present study of PHPT patients showed that, within 3 years after surgery, BMC of both cancellous and cortical bone areas had increased. At the same time, bone turnover decreased markedly, as judged from biochemical as well as histomorphometric data, but no changes were seen in trabecular bone structure. In cortical bone, the relative cortical width increased and the cortical porosity decreased.

Some new, simple and efficient stereological methods and their use in pathological research and diagnosis.

Stereology is a set of simple and efficient methods for quantitation of three-dimensional microscopic structures which is specifically tuned to provide reliable data from sections. Within the last few years, a number of new methods has been developed which are of special interest to pathologists. Methods for estimating the volume, surface area and length of any structure are described in this review. The principles on which stereology is based and the necessary sampling procedures are described and illustrated with examples. The necessary equipment, the measurements, and the calculations are invariably simple and easy.

Late nervous system disorders in cured malignant lymphoma: a clinical and neuropathological study.

A 53-year-old woman was treated for and cured of low grade malignant lymphoma, localized to the neck, by irradiation and chemotherapy. One year later she developed signs of damage to the spinal cord with slight paraparesis of the lower extremities, which remained stationary for seven years. Then, new and rapidly progressive central and peripheral neurological symptoms developed. About one year later the patient died. At autopsy a malignant glioma of the right temporal lobe and radiation damage to the spinal cord were found. Lymphocytic infiltrations in the peripheral nerves and muscles of the lower extremities were also seen. A severe neurogenic atrophy was present but no relapse of malignant lymphoma was found. Depressed immune defense is suggested to be the cause of the pathological changes of the nervous system in this case. The inflammation of the peripheral nerves might be due to activation of a latent virus infection.

Homicidal hanging masquerading as suicide.

Homicidal hanging is rare and presents special problems for the forensic pathologist. We report a case of homicide by hanging masquerading as suicide, in which the forensic evidence was of crucial importance. The victim was a 61 years old man, who was found in his house suspended by a rope around his neck. The autopsy showed fresh bruises due to blunt trauma. Moreover, a voluminous subdural haematoma with brain swelling was found which indicated that the victim was unconscious at the time of the hanging. An obvious vital reaction consisting of cutaneous bleeding in the ligature mark supported the opinion that the deceased was alive and not dead at the time of the hanging. The estimation of the time of death was of great importance, as only one other person could have been present at the estimated time interval. This person was later sentenced to prison for manslaughter.

Iatrogenic fractures of the hyoid bone and the thyroid cartilage. A case report.

A case of fracture of the hyoid bone and the thyroid cartilage is reported as a complication to resuscitation. This gave rise to many forensic as well as police investigations and stresses the importance of detailed information at medicolegal autopsies as regards the resuscitation carried out.

Sudden death in mitral valve prolapse: associated accessory atrioventricular pathways.

Sudden death in patients with mitral valve prolapse is a rare complication with a possible arrhythmic mechanism. We made a detailed postmortem examination of the conduction system in three patients with MVP who died suddenly. One patient who died in cardiac failure had a normal conduction system. The two other patients with no cardiac symptoms prior to death had both accessory atrioventricular pathways. These findings suggest a higher incidence than hitherto known of accessory bypass tracts in patients with MVP who die suddenly, and support the presumption of an arrhythmic cause of death.

Child abuse and neglect in Denmark: medico-legal aspects.

On the basis of material selected at the Institute of Forensic Medicine, Aarhus , from 1959 to 1980, we estimated the frequency of mortality from child abuse in Denmark to be 0.5 child deaths per million inhabitants per year. This figure is equal to that in Sweden and Norway but lower than most other countries. The study population included 53 abused children: 33 living children (18 boys and 15 girls) and 20 fatalities (11 boys and 9 girls). The social background of the children and abusers (32 men and 15 women) is described in connection with the juridical and social outcome of the cases. Cases with repetitive lesions were the most important evidence of child abuse. In cases with non-repetitive injuries, it was more difficult to prove that child abuse had taken place, but subdural hematoma and abdominal lesions with rupture of the duodenum or tearing of the mesenterium were very strong evidence. We think that a better handling of cases of child abuse or neglect could be obtained with extended collaboration between the social authorities, the police, the public prosecutor , the pediatrician and the forensic pathologist.