Double lamellae in trabecular osteons: towards a new method for age estimation by bone microscopy.

The study of anatomical ageing has a dual purpose in biological anthropology. On the one hand, it can provide insights into age associated changes in the body and thus widen the understanding of human senescence; and on the other hand it can provide means of estimation of age at death. This paper explores normal ageing in the pattern of remodelling of trabecular bone in humans. The material consists of necropsies of bone from the ilium of 25 males. A 1 cm2 prism extending from the outer to the inner surface of the iliac bone was removed from men who had died with no clinical signs of diseases, which would usually affect bone structure and metabolism. The samples were cut, and studied by light microscopy at a magnification of 100 x. New trabecular bone is formed in disk-shaped osteons with a clear double lamellar structure. In each sample, the number of double lamellae in a mean of 21 complete osteons was counted. The mean number of lamellae was taken as the measurement of interest. The log of the mean counts was found to regress linearly and with no evidence for heteroscedacity on age. The correlation between the two was high and negative (r=-0.83, p<0.001). The material is too limited to provide a useful basis for age estimation as such, but the study demonstrates the potential for palaeodemographic application of the method.

Plasma 25-hydroxyvitamin D and not 1,25-dihydroxyvitamin D is associated with parathyroid adenoma secretion in primary hyperparathyroidism: a cross-sectional study.

BACKGROUND: Primary hyperparathyroidism (PHPT) is associated with reduced plasma 25-hydroxyvitamin D (P-25OHD) and usually increased plasma 1alpha,25-dihydroxyvitamin D (P-1,25(OH)2D). Parathyroid tissue expresses the vitamin D receptor and it is thought that circulating 1,25(OH)2D participate in the regulation of parathyroid cell proliferation, differentiation and secretion. AIM: To investigate the relations between circulating levels of 1,25(OH)2D and 25OHD respectively and parathyroid adenoma weight (AW), plasma-parathyroid hormone (P-PTH) and PTH secretion expressed as P-PTH/AW. DESIGN: Cross-sectional study. MATERIAL: One hundred and seventy-one consecutive hypercalcaemic caucasian patients aged 19-87 years (median 63, 84% females) with surgically proven parathyroid adenoma. RESULTS: A weak positive correlation was found between P-25OHD and P-1,25(OH)2D (r=0.24, P<0.005). AW depended on sex and body mass index. Following adjustment, it was correlated positively to P-PTH, calcium (Ca) and alkaline phosphatase (AP) and inversely to plasma phosphate in a multiple regression model. AW was not associated with vitamin D metabolites. Preoperative P-PTH correlated positively to plasma levels of Ca and AP, but inversely to phosphate and 25OHD (P<0.001) levels. P-PTH was not associated with P-1,25(OH)2D (P=0.65). The P-PTH:AW ratio correlated inversely to P-25OHD (P<0.05), but showed no relations to plasma levels of Ca, phosphate or 1,25(OH)2D (P=0.22). CONCLUSION: In this material, low levels of 25OHD were related to higher levels of P-PTH and higher PTH:AW ratios in patients with PHPT suggesting that vitamin D deficiency increase PTH secretion activity. Neither PTH secretion nor AW was associated with circulating levels of 1,25(OH)2D.

Vitamin D status, seasonal variations, parathyroid adenoma weight and bone mineral density in primary hyperparathyroidism.

BACKGROUND: Primary hyperparathyroidism (PHPT) and vitamin D insufficiency are common conditions that can occur in combination. However, low plasma 25-hydroxyvitamin D (25OHD) may also enhance the risk of PHPT or modify disease severity. AIM: To compare the risk of vitamin D insufficiency and deficiency stratified by age, sex and season between PHPT patients and controls and to assess associations between plasma 25OHD and adenoma weight, biochemical variables, bone mineral density (BMD) and clinical complications. DESIGN: Cross-sectional study. MATERIAL: A total of 289 consecutive Caucasian patients with PHPT aged 65.9 (24-92) years, 289 sex-, age- and season-matched normocalcaemic controls and 187 healthy adult blood donors. PHPT diagnosis was confirmed in 214 by neck exploration. RESULTS: Vitamin D insufficiency (plasma 25OHD < 50 nmol/l) was observed in 81% of PHPT patients compared with 60% of sex- and age-matched controls (P < 0.001) and 35% of blood donors (P < 0.001). During summer, 77%vs. 53% (P < 0.001) and 4% (P < 0.001), respectively, had vitamin D insufficiency. Average plasma 25OHD was 41 (range 9-87) nmol/l among 27 PHPT patients compared with 87 (21-173) nmol/l (P < 0.001) among aged-matched blood donors. During winter, 86%vs. 66% (P < 0.001) and 71% (P < 0.05), respectively, had vitamin D insufficiency. Vitamin D deficiency (plasma 25OHD < 25 nmol/l) was observed in 33% of PHPT patients compared with 20% of age- and sex-matched controls (P < 0.001) and 13% of blood donors (P < 0.001). Both PHPT patients and controls showed seasonal variations in 25OHD related to the average number of sun hours, but values were lower in PHPT patients at all calendar months. In PHPT patients low plasma 25OHD was associated with higher plasma levels of calcium, PTH and alkaline phosphatase and with lower renal calcium excretion, femoral neck and forearm BMD. No association was found between plasma 25OHD and adenoma weight (total or divided into tertiles). There was a trend towards increased risk of osteoporotic fractures (P < 0.08) with low plasma 25OHD. CONCLUSION: Vitamin D insufficiency and deficiency are common findings in PHPT and occur more often than in a sex- and age-matched control group referred from general practice and in normal blood donors irrespective of season. Low plasma 25OHD levels are associated with an aggravated clinical presentation of PHPT but do not affect adenoma size.

MR imaging of the normal sacroiliac joint with correlation to histology.

OBJECTIVE: The microscopic study of the various components of joints provide a proper basis for understanding the nature of pathologic lesions to which they are subject and their imaging appearances. This study was designed to correlate MR imaging with a systematic histological study of the normal sacroiliac joint (SIJ), which to our knowledge is not available in the literature. DESIGN AND PATIENTS: Five male cadavers, aged 20 to 45 years, and seven male and seven female volunteers, aged 23 to 44 years, were investigated with oblique transaxial and coronal MR imaging of the SIJs. A variety of sequences including pre- and post-contrast T1 fat-saturated studies in the volunteers were used. Cryosectioning was performed in six SIJs of the five cadavers and compared with the MR images for the microscopic joint anatomy and assessed for the presence of abnormalities resembling those associated with sacroiliitis. RESULTS: Throughout the SIJ, the hyaline cartilage of the sacral bone and the proximal third of the hyaline iliac cartilage was strongly attached to the surrounding stabilizing ligaments, forming wide margins of fibrocartilage. In the distal one-third of the joint only, the margins of the iliac joint facet resemble that of a synovial joint, which include an inner capsule with synovial cells. The MR anatomy of the ventral and dorsal aspects of the SIJ was only adequately visualized at oblique transaxial MR imaging. No contrast enhancement occurred in the synovial tissue or in the cartilaginous joint space. The dorsal transition between the proximal 2/3 and distal 1/3 of the cartilaginous joint was at microscopy rich in anatomical and histological variants, including osseous clefts, cartilage and subchondral defects, and vascular connective tissue in the bone marrow. These were all recognized at oblique transaxial MR imaging and in coronal MR sectioning may resemble abnormalities. Otherwise, no erosions, bone marrow abnormalities, bone sclerosis or abnormal contrast enhancement occurred in the normal joints. CONCLUSIONS: The SIJ should be classified anatomically as a symphysis with some characteristics of a synovial joint being confined to the distal cartilaginous portion at the iliac side. Coronal MR imaging does not allow assessment of normal anatomy, variants or abnormalities of the ventral and dorsal margins of the cartilaginous SIJ.

Effects of long-term risedronate on bone quality and bone turnover in women with postmenopausal osteoporosis.

The effects of 3 years of oral risedronate treatment on bone quality and remodeling were assessed in women with postmenopausal osteoporosis. Transiliac bone biopsies were obtained at baseline and after treatment with placebo or risedronate 5 mg/day in 55 women (placebo, n = 27; risedronate 5 mg, n = 28); these pairs of samples allowed comparison of treatment effects vs. both baseline values and between treatment groups. A further 15 women (placebo, n = 6; risedronate 5 mg, n = 9) had measurements from a posttreatment biopsy, but not from a baseline biopsy. Samples were examined for qualitative changes (e.g., osteomalacia, peritrabecular fibrosis, and woven bone); no histological abnormalities were found to be associated with treatment. Among women with both baseline and posttreatment biopsies, risedronate-treated women experienced a moderate and expected reduction from baseline in bone turnover, which was reflected in mean decreases in mineralizing surface of 58% and in activation frequency of 47%. Histomorphometrical parameters indicated that bone formation rate decreased significantly from baseline with risedronate treatment, reflecting a decrease in bone turnover; bone mineralization was normal following treatment. Basic multicellular unit (BMU) balance tended to improve in the risedronate-treated women, whereas it tended to worsen in the placebo-treated women, although these changes were not statistically significant. There were no significant changes in structural parameters with treatment. The effects of 3 years of risedronate treatment on bone histology and histomorphometry reflect the antiresorptive mechanism of action, and are consistent with the antifracture efficacy and favorable bone safety profile demonstrated in large clinical trials.

Cancellous bone remodeling occurs in specialized compartments lined by cells expressing osteoblastic markers.

We describe a sinus, referred to as a bone remodeling compartment (BRC), which is intimately associated with cancellous bone remodeling. The compartment is lined on its marrow side by flattened cells and on its osseous side by the remodeling bone surface, resembling a roof of flattened cells covering the bone surface. The flat marrow lining cells are in continuity with the bone lining cells at the margins of the BRC. We examined a large number of diagnostic bone biopsy specimens received during recent years in the department. Furthermore, 10 patients (8 women and 2 men, median age 56 [40-69] years) with the high turnover disease of primary hyperparathyroidism who were treated with parathyroidectomy and followed for 3 years were included in the histomorphometric study. Bone samples for the immuno-enzyme staining were obtained from an amputated extremity of child. The total cancellous bone surface covered by BRC decreases by 50% (p < 0.05) following normalization of turnover and is paralleled by a similar 50% decrease in remodeling surface (p < 0.05). The entire eroded surface and two-thirds of the osteoid surface are covered by a BRC. BRC-covered uncompleted walls are 30% (p < 0.05) thinner than those without a BRC. This indicates that the BRC is invariably associated with the early phases of bone remodeling, that is, bone resorption, whereas it closes during the late part of bone formation. Immuno-enzyme staining shows that the flat marrow lining cells are positive for alkaline phosphatase, osteocalcin, and osteonectin, suggesting that they are bone cells. The first step in cancellous bone remodeling is thought to be the lining cells digesting the unmineralized matrix membrane followed by their disappearance and the arrival of the bone multicellular unit (BMU). We suggest that the lining cell barrier persists during bone remodeling; that the old lining cells become the marrow lining cells, allowing bone resorption and bone formation to proceed under a common roof of lining cells; that, at the end of bone formation, new bone lining cells derived from the flattened osteoblasts replace the marrow lining cells thereby closing the BRC; and that the two layers of lining cells eventually becomes a single layer. The integrity of the osteocyte-lining cell system is reestablished by the new generation of lining cells. The BRC most likely serves multiple purposes, including efficient exchange of matrix constituents and minerals, routing, monitoring, or modulating bone cell recruitment, and possibly the anatomical basis for the coupling of bone remodeling.

How many patients are needed? Variation and design considerations in bone histomorphometry.

In osteoporosis research, bone histomorphometry plays an important role in documenting the biological effects and possible side-effects of new drug treatments. To ensure that the study is properly scaled, it is important to be concerned with the risk of type II error; that is, the risk of failing to detect a real difference. We therefore calculated the necessary sample size in bone histomorphometric studies according to a specified difference of 15% between two groups. The calculations were based on variance components estimated from three different studies: women with a distal fracture of the forearm (n = 22); patients with pituitary insufficiency (n = 21); and patients with primary hyperparathyroidism (n = 21). Using a significance level of 0.05 and a risk of type II error of 0.20, the statistical power of two different designs was compared: a single biopsy design comparing the responses in two groups after the treatment; and a paired biopsy design in which individual differences (posttreatment minus baseline) were calculated before the comparison of the two groups. We found that the mineral apposition rate, wall thickness, and erosion depth are statistically powerful indices that, in the single biopsy design, require no more than n = 25 in each group to detect differences of 15% between the groups. Bone volume, erosion surface, osteoid surface, mineralizing surface, and activation frequency need group sizes of 100-600 individuals to find a 15% difference to be statistically significant. However, the effect of bisphosphonate treatment, for instance, is large enough to reduce the group size to 20 individuals concerning activation frequency. The remodeling balance reaches extreme group sizes of several thousand for a 15% difference to be statistically significant, but for a 5 microm (approximately 150%) improvement, about 100 individuals are required in the single biopsy design. An analysis of the components of variance showed that the variation between individuals is small and often negligible compared with the variation within individuals, and sample sizes needed for the paired biopsy design are therefore larger than those for the single biopsy design. In conclusion, the most cost-effective histomorphometric study design within a randomized clinical trial appears to be a single biopsy design comparing posttreatment biopsies with scaling performed according to the statistical power of the indices of interest.

Sentinel lymph node biopsy in breast cancer--the Aarhus experience.

Eighty patients, with newly diagnosed unifocal breast cancer and with no axillary metastases verified by ultrasonography, underwent sentinel lymph node (SLN) and subsequent axillary lymph node dissection. To identify the SLN, we used a combination of Tc-99m labelled colloid (Albures) and blue dye (Patent Blue V) injected peritumorally. Lymphoscintigraphy was not performed. The SLN was successfully identified in 78 out of 80 patients (97.5%); 43 patients (54%) were found to have metastatic disease. In 33 patients (77%) the SLN was the only node involved. No false-negative nodes were found, defined as SLNs that tested negative but with higher nodes that tested positive. If SLN biopsy is accepted as a routine procedure and when the exact indications are defined, the method described probably could be offered to the majority of breast cancer 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.

Growth hormone treatment in adults with adult-onset growth hormone deficiency increases iliac crest trabecular bone turnover: a 1-year, double-blind, randomized, placebo-controlled study.

The effects of growth hormone (GH) substitution on bone metabolism were evaluated by dynamic histomorphometry on iliac crest bone biopsies. Twenty-nine patients, aged 21-61 years (mean 45.5 years), with adult-onset GH deficiency (GHD) were randomized to receive subcutaneous injections with GH (2 IU/m2/day = 0.67 mg/m2/day) or placebo for 12 months. Serum insulin-like growth factor I (IGF-I) levels increased 263 +/- 98% (mean +/- SD) during GH treatment (p < 0.0001). In the GH group, osteoid surface increased during treatment from 11% (3-15%) (median [25-75 percentiles]) to 21% (10-27%) (p = 0.01) and mineralizing surface from 4% (1-8%) to 11%(7-16%) (p = 0.04). Moreover, erosion surface tended to increase in the GH group from 2% (1-3%) to 4% (3-5%) (p = 0.07). The quiescent surface decreased in the GH group from 87% (83-96%) to 74% (68-87%) (p = 0.01). The adjusted appositional rate, mineral apposition rate, bone formation rate, bone erosion rate, mineralization lag time, and osteoid thickness remained unchanged during treatment. Erosion depth showed a trend toward increase in the GH group (p = 0.09), whereas wall thickness was unchanged. Bone balance at the remodeling unit level and activation frequency were unchanged. At the tissue level, bone erosion rate increased significantly from 26% (17-36%)/year to 39% (23-72%)/year (p = 0.03). Similarly, the bone formation rate at the tissue level tended to increase, from 24% (15-31%)/year to 36% (17%-63%)%/year (p = 0.06). Finally, bone balance at the tissue level decreased significantly from 1% (-2-2%)/year to -5% (-13-1%)/year (p = 0.01). No significant difference in change was seen in the cancellous bone volume. We conclude that 12 months of GH substitution therapy increases trabecular bone turnover. Moreover, our data suggest that bone balance at the bone multicellular unit level is not changed to positive.

Differences in static cortical bone remodeling parameters in human mandible and iliac crest.

Knowledge of the baseline turnover characteristics, and of possible general and local factors influencing alveolar bone responses, is particularly important in the planning of oral rehabilitation. The conventional tool used to obtain information on bone turnover is the iliac crest biopsy, but it is not clear whether it mirrors the situation involving the jaws. The aim of this study was to compare static bone remodeling parameters in the mandible and in the iliac crest to obtain baseline values for the mandible and to test the hypothesis of site specificity of bone remodeling. Bone specimens were obtained from 50 subjects (mean age 64 +/- 17) at autopsy. Three sites were sampled: iliac crest; jaw angle; and foramen mentalis area. In addition, occlusal status was recorded. On undecalcified thin sections, cortical porosity (Ct.Po), eroded sites (ESi), formative sites (FSi), osteonal diameter (On.Dm), Haversian canal diameter (H.Ca.Dm), and wall width (W.Wi) were measured. Ct.Po in the jaw angle and in the foramen mentalis area was lower (48% and 50%, respectively) than in the iliac crest, as was ESi and FSi (80% in the jaw angle and 74% in the foramen mentalis area). In the foramen mentalis area, Ct.Po was greater in subjects with occlusion. On.Dm, H.Ca.Dm, and W.Wi were significantly larger and mutually correlated within the mandible, whereas no correlation was found between mandibular sites and iliac crest. Static cortical bone remodeling parameters are different in the mandible and the iliac crest, thus confirming the hypothesis of site specificity of bone remodeling. Within the mandible, the parameters were correlated, whereas there was no correlation between the mandible and the iliac crest. This could be ascribed to the different functional demands to the mandible and the iliac crest, which was also reflected in the observed influence of functional occlusion on bone remodeling in the mandible. It can thus be concluded that bone reaction to dental intervention is more dependent on the local environment than on general bone turnover as reflected by the iliac crest.

Primary hyperparathyroidism: effect of parathyroidectomy on regional bone mineral density in Danish patients: a three-year follow-up study.

Changes in skeletal remodeling (biochemical bone markers) and regional bone mineral density (spine, hip, and forearm bone mineral density [BMD]) were observed for 3 years in 20 patients (15 women and 5 men; age 54 +/- 11 years, range 29-69 years) after successful surgery for primary hyperparathyroidism (PHPT). Fifteen PHPT patients were compared with 15 normal controls who were exactly matched with respect to age, gender, and menopausal status (10 women and 5 men; age 53 +/- 12 years, range 29-65 years [PHPT] and 29-66 years [controls]). All bone markers (serum osteocalcin, bone alkaline phosphatase, and type I collagen telopeptide [ICTP], and urinary hydroxyproline and NTx/creatinine ratio) declined significantly and reached normal levels within 6 months. No major changes took place during the remaining 2.5 years, apart from urine hydroxyproline, which disclosed a small peak around 12 months with a further decline towards study end (p < 0.05). Bone mineral density increased significantly in all regions (p < 0.001). At all locations, except the intertrochanteric region of the hip, the increase continued from 6 months until study end (p < 0.05). The increase in BMD was unequally distributed among regions (p < 0.001). The increase at the proximal forearm was less than in the spine (p < 0.05), the trochanteric region of the hip (p < 0.05), and the distal forearm (p < 0.05). No difference in BMD increase was observed between men, and pre- and postmenopausal women. Compared with the matched control group, PHPT patients had significantly lower BMD at baseline in the proximal (p < 0.02) and distal (p < 0.05) forearm. Furthermore, during the 3-year follow-up period, the PHPT patients showed a significant increase in BMD compared with controls in the spine (p < 0.005), the trochanteric and intertrochanteric regions of the hip (p < 0.005 and p < 0.05, respectively), and the distal forearm (p < 0.005). In conclusion, bone remodeling is normalized within the first 6 months after successful parathyroid surgery, with no major changes during the following 2.5 years. Bone mineral density increases at both cancellous and cortical sites, but in predominantly cortical bone, the recovery in BMD is less than in cancellous bone-rich areas.

Primary hyperparathyroidism: whole-body bone mineral density in surgically treated Danish patients: a three-year follow-up study.

Whole-body bone mineral density (BMD) and body composition were measured before surgery in 25 patients (20 women and 5 men, aged 53 +/- 13 years, range 26-73 years) with mild to moderate primary hyperparathyroidism (PHPT) and compared with 25 controls exactly matched with respect to age, gender, and menopausal status. Fifteen pairs of matched patients and controls were reexamined 3 years later (5 men and 10 women, aged 53 +/- 12 years in both groups). In the untreated PHPT patients, whole-body BMD was 95.4% +/- 10.5% (SD) of control BMD (p < 0.05). Body weight and height, body mass index, whole-body fat mass, and lean body mass did not differ significantly between the groups. Relative to values in matched controls, whole-body bone mineral content (BMC) and BMD increased by 4.4% and 3.0%, respectively, in PHPT patients (p < 0.005) during the 3-year follow-up. Neither whole-body BMC nor BMD differed between patients and controls after the 3-year follow-up. A positive correlation was observed between initial serum calcium levels and the 3-year increase in whole-body BMD (r(s) = 0.645, p < 0.01). Baseline serum osteocalcin, serum pyridinoline crosslinked telopeptide of Type I collagen and several histomorphometric indices of trabecular bone turnover (eroded and labeled surfaces, bone formation rate, and activation frequency) also correlated positively with the subsequent increase in whole-body BMD. Six patients disclosed transient postoperative secondary hyperparathyroidism, probably due to hungry bones. Four of these patients completed 3 years of follow-up and had higher increases in whole-body BMD than the remaining normo-parathyroid patients (7.9% +/- 4.5%, range 4.3-14.3% versus 1.9% +/- 2.1%, p < 0.01). It is concluded that Danish patients with mild to moderate PHPT only reveal small reductions in whole-body mineral density. Furthermore, within 3 years after parathyroid surgery, most of the lost bone mineral is regained even in patients with initial high bone turnover. Finally, PHPT in these patients is not associated with substantial changes in body compositions.

Missing observations in bone histomorphometry on osteoporosis: implications and suggestions for an approach.

Crucial bone histomorphometric indices, i.e., turnover-related indices, are based on tetracycline double labeling. However, these indices are particularly exposed to loss of information because of missing readings on double labels. If the failure to make the observation is related to its magnitude, then selection bias may invalidate the conclusions. Therefore, ignoring missing double labels may lead to a selection of high-turnover patients. The aim of this study was to analyze the dimension and the impact of excluding iliac crest bone biopsies with missing readings in women with spinal crush fracture osteoporosis (n = 158, median 68 years, range 49-80 years). Furthermore, two different lower limits of the mineral apposition rate (MAR) were examined to explore their usefulness as a biological minimum that can be used for cases with missing readings, i.e., recoding of missing values. The average MAR (calculated as the mean of all interlabel widths measured in each individual) shows a lower limit of 0.3 microm/day, suggesting an apparent minimum for the interlabel width (Ir.L.Wi) of 3-4 microm. Identifying the smallest interlabel width measured in each individual and calculating the minimal MAR shows that 77% of the minimal MAR values are below 0.3 microm/day and reach a minimum of 0.1 microm/day, corresponding to an interlabel width of about 1 microm. Therefore, the minimal MAR presents a biological minimum of 0.1 microm/day. This value is used for our recoding: if no labels are sampled (2% of our population), Ir.L.Wi is assigned the value 0; if none or an insufficient number of double labels are sampled (29% of our population), then Ir.L.Wi is assigned the value 1 microm. Excluding cases with missing readings on any dependent variable increases the mineralizing surface (MS/BS) by 60% (2p < 0.01); other indices show no significant change. The suggested recoding decreases the average MAR by 4% (2p < 0.01), prolongs the remodeling period by 19% (2p < 0.01), and tends to decrease the activation frequency (2p = 0.09). Furthermore, the number of excluded biopsies tends to be larger among the older (2p = 0.09) and more severely osteopenic individuals (2p = 0.09). We conclude that ignoring missing double labels leads to selection bias; therefore, specific measures such as recoding procedures are needed to allow proper representation of low turnover patients. There is also a risk of bias caused by the exclusion of the older, osteopenic patients in bone histomorphometric osteoporosis trials.

Primary hyperparathyroidism: short-term changes in bone remodeling and bone mineral density following parathyroidectomy.

Changes in bone remodeling and bone mineral density were observed during a period of 6 months after surgery in 24 patients with primary hyperparathyroidism (20 women and 4 men; age 54+/-12 years, range 26-69 years). All bone markers declined significantly within the 6 month follow-up period, but the time course for changes in renal N-terminal telopeptide of type 1 collagen (NTx) excretion differed from those of the other markers by a steep and significant reduction (p < 0.05) after less than 1 month. During the 6 month period, bone mineral density (BMD) increased significantly at all sites measured (p < 0.05) apart from the femoral neck and the proximal and midforearm. The greatest increase of 4.2% was observed in the trochanteric region (p < 0.001). The increase in BMD in spine, trochanteric, and intertrochanteric regions of the hip correlated inversely with baseline forearm BMD values (p < 0.05). Baseline bone markers (serum alkaline phosphatase [AP], serum bone AP, serum pyridinoline crosslinked telopeptide of type 1 collagen, urinary hydroxyproline, urinary osteocalcin), as well as baseline histomorphometric indices of bone turnover (eroded and labeled surface, bone formation rate, activation frequency, and cortical porosity) were positively correlated with changes in spinal BMD over 6 months (p < 0.05). It was concluded that, within 6 months after parathyroidectomy, patients with primary hyperparathyroidism obtain normalization of bone remodeling and a substantial increase in bone mineral density in regions rich in cancellous bone but no significant changes in regions with predominantly cortical bone.

An endocytic pathway essential for renal uptake and activation of the steroid 25-(OH) vitamin D3.

Steroid hormones may enter cells by diffusion through the plasma membrane. However, we demonstrate here that some steroid hormones are taken up by receptor-mediated endocytosis of steroid-carrier complexes. We show that 25-(OH) vitamin D3 in complex with its plasma carrier, the vitamin D-binding protein, is filtered through the glomerulus and reabsorbed in the proximal tubules by the endocytic receptor megalin. Endocytosis is required to preserve 25-(OH) vitamin D3 and to deliver to the cells the precursor for generation of 1,25-(OH)2 vitamin D3, a regulator of the calcium metabolism. Megalin-/- mice are unable to retrieve the steroid from the glomerular filtrate and develop vitamin D deficiency and bone disease.

A histomorphometric study of haematological disorders with respect to marrow fibrosis and osteosclerosis.

A retrospective investigation of 75 EDTA-decalcified Jamshidi biopsies collected over a 2-year period at Aarhus University Hospital was performed. The biopsies originated from 75 patients suffering from idiopathic myelofibrosis, other chronic myeloproliferative disorders, or other conditions with known associations with bone marrow fibrosis. The relative volumes of trabecular and woven bone, as well as haematopoietic and non-haematopoietic tissue, were estimated histomorphometrically. The degree of fibrosis was estimated semiquantitatively. Finally, the thickness of trabecular osteons was estimated from the number of lamellae by counting. Patients with idiopathic myelofibrosis had statistically significantly more bone tissue than the other groups of patients. The osteosclerosis was primarily due to woven bone. Larger cancellous osteons also suggested a positive balance in bone remodelling. The amount of bone tissue showed furthermore a statistically significant increase through the groups of polycythaemia vera, essential thrombocythaemia, chronic myelogenous leukaemia and idiopathic myelofibrosis. Parallel to the increase in the amount of bone, an increase in the degree of marrow fibrosis was detected. The positive correlation between the amount of bone and the degree of marrow fibrosis was statistically highly significant (p=0.0008).

MIB-1 expression in breast carcinomas with medullary features. An immunohistological study including correlations with p53 and bcl-2.

Typical medullary carcinoma (TMC) is usually considered to have a more favourable prognosis than other types of infiltrating breast carcinomas. This is a biological paradox, since its clinical behaviour is not in agreement with its anaplastic morphology and high mitotic rate. It should be remembered that neoplastic growth reflects cell production minus cell loss, the latter being achieved by apoptosis. At present, bcl-2 oncogene (apoptosis inhibitor) and p53 gene are assumed to be involved in the regulation of cell death and tumour proliferation. Sixty breast carcinomas, initially indexed as medullary carcinomas, were re-classified using the diagnostic criteria given by Ridolfi. This review yielded 13 typical (TMC), 24 atypical (AMC), and 23 non-medullary carcinomas (NMC). Following antigen retrieval by microwave treatment, immunohistochemical analyses, using MIB-1, p53 and bcl-2 monoclonal antibodies were performed on serial sections from formalin-fixed, paraffin-embedded specimens. TMC revealed the highest incidence of intense p53 positivity, and the highest mean MIB-1 index, and absence of the apoptosis-inhibitor protein bcl-2. These results suggest the presence of a higher overall cell turnover in TMC than in AMC and NMC. Increased apoptosis balancing the increased cell proliferation might be among the possible explanations for the more favourable prognosis in TMC.

Primary hyperparathyroidism: biochemical markers and bone mineral density at multiple skeletal sites in Danish patients.

Biochemical bone markers and bone mineral density (BMD) in spine, hip, and forearm were measured, before surgery, in 30 patients with mild to moderate primary hyperparathyroidism (PHP) (25 women and 5 men; mean age 54 +/- 12 years, range 26-73 years) and compared with normal controls. A group of 291 healthy adults (181 women and 110 men) served as controls for BMD. A smaller group of 30 normal individuals (25 women and 5 men; mean age 54 +/- 12 years; range 26-74 years) were used as matched normal controls. Parameters of bone formation (s-osteocalcin, s-alkaline phosphatase activity, and s-bone isoenzyme alkaline phosphatase activity) and bone resorption (s-type-1 collagen telopeptide) were considerably increased in patients with PHP compared with normal controls (p < 0.01 for all parameters). BMD was found to be reduced in the hip (trochanteric: 95.1 +/- 14.7% of expected, p < 0.05; intertrochanteric: 95.2 +/- 13.8% of expected, p < 0.05), and the forearm (proximal: 93.3 +/- 12.2% of expected, p < 0.05; mid: 91.8 +/- 11.6% of expected, p < 0.001; distal: 90.7 +/- 13.1% of expected, p < 0.001). Spine BMD was found significantly reduced in premenopausal (87.8 +/- 7.6% of expected, p < 0.05) but not in postmenopausal patients, and although normal women showed a decrease in spinal BMD with increasing age this was not found in the PHP women. Forearm BMD was reduced in both pre- and postmenopausal patients (distal forearm: 86.7 +/- 12.2% of expected, p < 0.05; 87.6 +/- 12.1% of expected, p < 0.01, respectively). It was concluded that Danish patients with mild or moderate PHP have only small reductions in BMD. The bone loss is mainly found in the appendicular skeleton.

Prognostic comparison of three classifications for medullary carcinomas of the breast.

The aim of this study was to make prognostic comparisons between the modified scheme of Pedersen et al. the definitions of Tavassoli and the Ridolfi criteria for medullary carcinomas. Sixty breast carcinomas primarily diagnosed as medullary carcinomas were reclassified into typical medullary carcinoma (TMC), atypical medullary carcinoma (AMC) and non-medullary carcinoma (NMC) according to the three classifications. The Ridolfi classification proved to be superior to the two other schemes in discriminating survival differences between the three groups TMC, AMC and NMC. All 13 patients with TMC are still alive indicating an excellent prognosis, while 29% and 39% of the 47 patients in the AMC and NMC category, respectively, have died of their disease. In the simplified system of Pedersen et al. the survival at 10 years for TMC patients decreased to 75% and no significant survival difference between the three groups could be demonstrated. As the prognosis for AMC proved to be worse compared to TMC and in fact was similar to NMC with values of 43% at 10 years in the Ridolfi classification, we find no reasons to maintain this category. We conclude that as long as no alternative and more easily applicable diagnostic method exists, pathologists should still apply the Ridolfi criteria on these tumours with medullary features leaving two diagnostic possibilities: TMC or NMC (i.e. poorly differentiated ductal carcinoma). Only lesions that fulfil all six criteria without any doubt should be diagnosed as TMC, thus avoiding overdiagnosis and a resulting risk of undertreatment.