One Year of Transgenic Overexpression of Osteoprotegerin in Rats Suppressed Bone Resorption and Increased Vertebral Bone Volume,Density, and Strength

RANKL is an essential mediator of bone resorption, and its activity is inhibited by osteoprotegerin (OPG). Transgenic (Tg) rats were engineered to continuously overexpress OPG to study the effects of continuous long‐term RANKL inhibition on bone volume, density, and strength. Lumbar vertebrae, femurs, and blood were obtained from 1‐yr‐old female OPG‐Tg rats (n = 32) and from age‐matched wildtype (WT) controls (n = 23). OPG‐Tg rats had significantly greater serum OPG (up to 260‐fold) and significantly lower serum TRACP5b and osteocalcin compared with WT controls. Vertebral histomorphometry showed significant reductions in osteoclasts and bone turnover parameters in OPG‐Tg rats versus WT controls, and these reductions were associated with significantly greater peak load in vertebrae tested through compression. No apparent differences in bone material properties were observed in OPG‐Tg rat vertebrae, based on their unchanged intrinsic strength parameters and their normal linear relationship between vertebral bone mass and strength. Femurs from OPG‐Tg rats were of normal length but showed mild osteopetrotic changes, including reduced periosteal perimeter (?6%) and an associated reduction in bending strength. Serum OPG levels in WT rats showed no correlations with any measured parameter of bone turnover, mass, or strength, whereas the supraphysiological serum OPG levels in OPG‐Tg rats correlated negatively with bone turnover parameters and positively with vertebral bone mass and strength parameters. In summary, low bone turnover after 1 yr of OPG overexpression in rats was associated with increased vertebral bone mass and proportional increases in bone strength, with no evidence for deleterious effects on vertebral material properties.     

Genetic Variations in Bone Density, Histomorphometry, and Strength in Mice

The purpose of this study was to assess breed-related differences in bone histomorphometry, bone biomechanics, and serum biochemistry in three mouse breeds shown to differ in bone mineral density (BMD) (as measured by DXA) and bone mineral content (BMC). Femurs, tibiae, and sera were collected from 16-week-old C3H/HeJ {C3H}, C57BL/6J {BL6}, and DBA/2J {DBA}mice (n = 12/breed). Data collected included BMC and BMD (femora), histomorphometry of cancellous (distal femur) and cortical bone (diaphyseal tibiae and femora), bone strength (femora), and serum alkaline phosphatase (ALP). Consistent with previous reports, BMC and BMD were higher in C3H than in BL6 or DBA mice. The higher BMD in the C3H breed was associated with greater cancellous bone volume, cortical bone area, periosteal bone formation rate, biomechanical strength, and serum ALP. However, mid-diaphyseal total femoral and tibial cross-sectional area and moment of inertia were greatest in BL6, intermediate in C3H, and lowest in DBA mice. The specific distribution of cortical bone in C3H, BL6, DBA mice represents a difference in adaptive response to similar mechanical loads in these breeds. This difference in adaptive response may be intrinsic to the adaptive mechanism, or may be intrinsic to the bone tissue material properties. In either case, the bone-adaptive response to ordinary mechanical loads in the BL6 mice yields bones of lower mechanical efficiency (less stiffness per unit mass of bone tissue) and does not adapt as well as that of the C3H mice where the final product is a bone with greater resistance to bending under load. We suggest that the size, shape, and BMD of the bone are a result of breed-specific genetically regulated cellular mechanisms. Compared with the C3H mice, the lower BMD in BL6 mice is associated with long bones that are weaker because the larger cross-sectional area fails to compensate completely for their lower BMD and BMC. Received: 16 November 1999 / Accepted: 19 April 2000 / Online publication: 27 July 2000     

Bone Quality and Bone Strength in BXH Recombinant Inbred Mice

The relationship between bone quality and strength was studied in 11 BXH recombinant inbred (RI) strains of mice. The bone quality parameters studied were bone mineralization, microhardness, architecture, and connectivity. Previous studies have demonstrated considerable variability in bone density, biomechanical properties, and microstructure among inbred strains of mice. In particular, C3H/HeJ (C3H) mice exhibit thicker femoral and vertebral cortices and fewer trabeculae in the vertebral body compared with C57BL/6J (B6) mice, despite having similar vertebral bone strength. A set of RI mouse strains has been generated from B6 and C3H (denoted BXH) in an attempt to isolate genetic regulation of numerous traits, including bone. The objective of this study was to investigate relationships among bone quality and bone strength in femurs and vertebrae among BXH RI mice. The study involved 11 BXH RI strains of female mice (n = 5−7) as well as the B6 and C3H progenitor strains. Parameters contributing to bone quality were evaluated, including BMD, bone mineralization, microhardness, architecture, and connectivity. There was a strong correlation between femoral and vertebral BMD in all strains (P < 0.001) except in BXH-9 and -10 (P < 0.001). Within the vertebrae, cortical bone was more mineralized than trabecular bone, and a strong correlation existed between the two (P < 0.001). However, cortical microhardness did not differ from trabecular microhardness. Cortical bone was more mineralized in the femur than in the vertebrae and significantly harder, by 30%. There was a wide range in trabecular connectivity, architecture, and femur geometry among BXH RI strains. BMD explained 43% of vertebral bone strength but only 11% of femoral bone strength. Trabecular connectivity explained an additional 8% of vertebral strength, while mineralization and femur geometry explained 7% and 50% of femoral strength, respectively. Different bone quality parameters had varying influences on bone mechanical properties, depending on bone site. BMD may play a larger role in explaining bone strength in the vertebrae than in the femur. Moreover, cortical bone in the femur is harder than in vertebrae. The control of cortical bone material properties may be site-dependent.     

Changes in Osteoprotegerin/RANKL System, Bone Mineral Density, and Bone Biochemicals Markers in Patients with Recent Spinal Cord Injury

This study analyzed the temporal and regional variations in bone loss and explored bone cell activities via biochemical markers during an extended follow-up in patients with spinal cord injury (SCI). In parallel, the possible role of the osteoprotegerin (OPG)/RANKL system in disuse osteoporosis was investigated. Seven male patients with acute and complete SCI (31.3 ± 9.5 years) and 12 able-bodied (AB) men (26.9 ± 4.2 years) participated in the study. Measurements were performed 16, 24, 36, 48, and 71 weeks after injury. At week 16, marked calcium homeostasis disturbance and a concomitant increase in bone resorption markers were observed, reflecting an intense bone degradation process. Resorption activity decreased continuously with time. Contrasting with the great rise in the resorption markers, the bone formation markers showed little variation. During the period of investigation, a loss in bone mineral density (BMD) was demonstrated for the total body (−4.3%), pelvis (−15.7%) and lower limbs (−15.2%), whereas BMD did not change at the lumbar spine, upper limbs, or skull. At all stages, SCI patients had lower serum RANKL levels and higher serum OPG levels than did AB controls, but no significant variation with time was observed for either cytokine. These findings suggest that bone resorption persisted long after SCI and specifically affected BMD at sublesional sites. The marked modification of serum OPG/RANKL levels in SCI patients suggests that this system is affected, in disuse osteoporosis. However, the precise biologic role of the OPG/RANKL system in the bone tissue of SCI patients has yet to be determined.     

脊髓损伤对大鼠骨转换及骨密度的影响

目的：探讨脊髓损伤（spinal cord injury,SCI）对大鼠骨转换及骨密度的影响。方法：60只3月龄SD大鼠均分为SCI组与对照组。SCI组于T10处完全横断脊髓;对照组仅行椎板切除术。术后1、3、6周时处死动物测血清钙（Ca）、磷（P）、碱性磷酸酶（ALP）、尿钙、磷、肌酊（Cr）以及股骨、股骨骨密度（bone mineral density,BMD）。:桂花 血钙、尿钙、尿钙／肌酝在伤后不同时间段均升高;血ALP在伤后1周时显著下降,3、6周时恢复正常。胫骨近端,股骨远、近端的BMD在6周时较对照组下降且差异显著。结论：SCI大鼠可见明显的骨转换增高以及破骨活性增强,其生化、骨密度的改变与人体有较好的相关性。SD大鼠可用于评价SCI后骨代谢改变,具可以作为SCI后骨质疏松的模型。     

Risedronate Preserves Bone Architecture in Early Postmenopausal Women In 1 Year as Measured by Three-Dimensional Microcomputed Tomography

Risedronate reduces the risk of vertebral fractures by up to 70% within the first year of treatment. Increases in bone mineral density or decreases in bone turnover markers explain only a portion of the anti-fracture effect, suggesting that other factors, such as changes in trabecular bone architecture, also play a role. Our objective was to determine the effects of risedronate on bone architecture by analyzing iliac crest bone biopsy specimens using three-dimensional microcomputed tomography (3-D µCT). Biopsy specimens were obtained at baseline and after 1 year of treatment from women enrolled in a double-blind, placebo-controlled study of risedronate 5 mg daily for the prevention of early postmenopausal bone loss. Trabecular architecture deteriorated in the placebo group (n = 12), as indicated by a 20.3% decrease in bone volume (25.1% vs. 20.0%, P = 0.034), a 13.5% decrease in trabecular number (1.649 vs. 1.426 mm^–1, P = 0.052), a 13.1% increase in trabecular separation (605 vs. 684 µm, P = 0.056), and an 86.2% increase in marrow star volume (3.251 vs. 6.053 mm³, P = 0.040) compared with baseline values. These changes in architectural parameters occurred in the presence of a concomitant decrease from baseline in lumbar spine bone mineral density (–3.3%, P = 0.002), as measured by dual energy x-ray absorptiometry. There was no statistically significant (P < 0.05) deterioration in the risedronate-treated group (n = 14) over the 1-year treatment period. Comparing the actual changes between the two groups, the placebo group experienced decreases in bone volume (placebo, –5.1%; risedronate, +3.5%; P = 0.011), trabecular thickness (placebo, –20 µm; risedronate, +23 µm; P = 0.032), and trabecular number (placebo, –0.223 mm^–1; risedronate, +0.099 mm^–1; P = 0.010), and increases in percent plate (placebo, +2.79%; risedronate, –3.23%; P = 0.018), trabecular separation (placebo, +79 µm; risedronate, –46 µm; P = 0.010) and marrow star volume (placebo, +2.80 mm³ ; risedronate, –2.08mm³; P = 0.036), compared with the risedronate group. These data demonstrate that trabecular architecture deteriorated significantly in this cohort of early postmenopausal women, and that this deterioration was prevented by risedronate. Although there is no direct link in this study between fracture and preservation of architecture, it is reasonable to infer that the preservation of bone architecture may play a role in risedronates anti-fracture efficacy.     

Hypothalamic Suppression Decreases Bone Strength Before and After Puberty in a Rat Model

The incidence of menstrual irregularities, both primary and secondary amenorrhea, has been reported to be as high as 60%, with the highest incidence in younger athletes, suggesting possible adverse effects on bone development. It was hypothesized that in a rat model, suppressed hypothalamic activity via a gonadotropin-releasing hormone antagonist (GnRH-a) before onset of puberty would result in a relatively larger bone strength deficit compared with suppression after puberty. Hypothalamic suppression was achieved by providing GnRH injections. Animals received injections for 25 days either before puberty (pre group) (age 23–46 days) or after puberty (post group) (age 65–90 days). Body weights and uterine weights were measured. Serum estradiol was assayed. Mechanical strength of the right femora and histomorphometry of the left femur were measured. Suppression of the hypothalamic–pituitary–gonadal axis was confirmed by significant atrophy of uterine tissue and suppressed estradiol levels. The peak moment was significantly lower in the pre and post GnRH-a groups compared with control. The percentage difference of the average peak moment and stiffness values from the respective age-matched control groups yielded a greater percentage difference in the pre group. The cortical area was less in the GnRH-a-treated groups, but no significant difference between the relative deficits between pre and post groups were found. Hypothalimic–pituitary–gonadal axis suppression before puberty resulted in a significantly larger deficit in mechanical strength compared with postpubertal animals. The time before puberty may represent a time when skeletal strength is more compromised. Women experience both primary and secondary amenorrhea; however, the treatment may need to be different for each condition.     

Osteogenesis Imperfecta: Bone Turnover, Bone Density, and Ultrasound Parameters

We studied 21 patients (11 men and 10 women) with osteogenesis imperfecta (OI) and 21 age- and sex-matched controls. In all patients we measured serum levels of total alkaline phosphatase (ALP), type I procollagen carboxy-terminal propeptide (PICP), osteocalcin (BGP), urinary excretion of hydroxyproline (HOP/Cr), and pyridinoline crosslinks (Pyr/Cr). Bone mineral density was measured at the distal radius (BMD-R) and at the lumbar spine (BMD-LS) by dual X-ray absorptiometry (DXA). Ultrasound parameters were also performed at the calcaneous with the Achilles device and at the phalanxes with DBM Sonic 1200. A significant reduction (P < 0.001) in BMD and in ultrasound parameters was found in OI patients compared with normals. PICP was significantly reduced in the OI patients compared with controls (P < 0.001); other markers of bone turnover were higher in OI than in controls, but the difference did not reach the statistical significance. A significant correlation (P < 0.05) was found between PICP and BMD at the lumbar spine and between PICP and ultrasound parameters at the calcaneous. On the basis of our data, we conclude that patients with OI show low values of BMD and ultrasound parameters; therefore in these patients, not only is bone mass disturbed but also bone quality. The reduced levels of PICP in OI patients confirm that most OI patients have defects in collagen I biosynthesis. These defects may contribute to the fragility of OI bone by interfering with complete mineralization and/or normal tissue structure. PICP may be considered a useful marker in the clinical management of OI. Received: 26 March 1998 / Accepted: 15 January 1999     

Colles' Fracture of the Wrist as an Indicator of Underlying Osteoporosis in Postmenopausal Women: A Prospective Study of Bone Mineral Density and Bone Turnover Rate

Colles' fracture has been shown to be associated with an increased risk of hip fracture. The incidence of low bone mineral density (BMD) and high bone turnover in such patients is uncertain. The aim of this study was to prospectively assess BMD and bone turnover in a cohort of consecutive postmenopausal Colles' fracture patients. BMD (spine, hip and contra-lateral radius) was measured by dual-energy X-ray absorptiometry (DXA) within 2 weeks of fracture. Bone turnover was assessed within 4 days by measurement of serum osteocalcin, total alkaline phosphatase (TALP), bone-specific alkaline phosphatase (BSAP) and urine hydroxyproline. We recruited 106 (71%) of 149 consecutive patients. Fifty-one per cent of subjects had a history of previous fracture, and 25% a past history of wrist, hip or vertebral body fracture. The incidence of osteoporosis was 21%, 42% and 22% at the spine, hip and radius respectively. Fifty per cent of subjects had osteoporosis of at least one of these sites. When compared with the values expected for their age the patients were found to have higher BMD than expected at the spine, and slightly lower BMD at the hip and distal radius. Patients aged 65 years or less had lower hip BMD than expected from the age-matched normal range (p < 0.01). Osteocalcin and TALP levels did not differ from the normal ranges, but BSAP and hydroxyproline levels were significantly elevated (p < 0.001), within 37% and 25% of patients having levels above the respective normal ranges. We conclude that osteoporosis is common in patients with Colles' fracture; however, in older patients BMD is not lower than would be expected in the normal population. In patients aged 65 years or less BMD is lower than expected at the hip. Bone turnover rate is high in many such patients. Intervention to prevent future fracture would be appropriate in women aged 65 years or less with Colles' fracture.     

Bone Volume Fraction Explains the Variation in Strength and Stiffness of Cancellous Bone Affected by Metastatic Cancer and Osteoporosis

Preventing nontraumatic fractures in millions of patients with osteoporosis or metastatic cancer may significantly reduce the associated morbidity and reduce health-care expenditures incurred by these fractures. Predicting fracture occurrence requires an accurate understanding of the relationship between bone structure and the mechanical properties governing bone fracture that can be readily measured. The aim of this study was to test the hypothesis that a single analytic relationship with either bone tissue mineral density or bone volume fraction (BV/TV) as independent variables could predict the strength and stiffness of normal and pathologic cancellous bone affected by osteoporosis or metastatic cancer. After obtaining institutional review board approval and informed consent, 15 patients underwent excisional biopsy of metastatic prostate, breast, lung, ovarian, or colon cancer from the spine and/or femur to obtain 41 metastatic cancer specimens. In addition, 96 noncancer specimens were excised from 43 age- and site-matched cadavers. All specimens were imaged using micro-computed tomography (micro-CT) and backscatter emission imaging and tested mechanically by uniaxial compression and nanoindentation. The minimum BV/TV, measured using quantitative micro-CT, accounted for 84% of the variation in bone stiffness and strength for all cancellous bone specimens. While relationships relating bone density to strength and stiffness have been derived empirically for normal and osteoporotic bone, these relationships have not been applied to skeletal metastases. This simple analytic relationship will facilitate large-scale screening and prediction of fracture risk for normal and pathologic cancellous bone using clinical CT systems to determine the load capacity of bones altered by metastatic cancer, osteoporosis, or both.     

l-Carnitine Fumarate and Isovaleryl-l-Carnitine Fumarate Accelerate the Recovery of Bone Volume/Total Volume Ratio after Experimetally Induced Osteoporosis in Pregnant Mice

Anabolic skeletal agents have recently broadened the therapeutic options for osteoporosis by directly stimulating bone formation and improving bone turnover, bone density, bone size, and bone microarchitecture. We recently demonstrated that two new l-carnitine derivatives, l-carnitine fumarate (LC) and isovaleryl-l-carnitine fumarate (Iso-V-LC), stimulated osteoblast proliferation and differentiation. We here investigated, by histomorphometry in a mouse model of osteoporosis, the impact of these compounds on the repair of trabecular bone and the osteoblast involvement in this process. Fifty-nine inbred adult female CD1 mice in pregnancy were assigned to four treatment groups: (1) controls, mice fed a standard normocalcemic pre- and postpartal diet; (2) Hypo, mice fed a low-calcium isocaloric prepartal diet and a standard postpartal diet; (3) LC, mice fed a group 2-type diet supplemented post-partum with LC; (4) Iso-V-LC, mice fed a group 2-type diet supplemented post-partum with Iso-V-LC. Bone volume/total volume ratio (BV/TV), bone perimeter, osteoblast surface/bone surface, and osteoblast number/bone surface were measured from sections of L3 and L4 vertebral bodies obtained from animals killed on the day of delivery (controls and Hypo) and on days 7, 14, and 21 after delivery (all groups). BV/TV and all osteoblast-based indexes were significantly higher in LC and Iso-V-LC than in Hypo mice at each time point, and Iso-V-LC at the end of the treatment attained levels observed in controls. In conclusion, Iso-V-LC and, to a lesser extent, LC accelerated the recovery of normal BV/TV level after a hypocalcemic diet.     

Non-association of Estrogen Receptor Genotypes with Bone Mineral Density and Bone Turnover in Korean Pre-, Peri-, and Postmenopausal Women

Estrogen is known to play a critical role in both skeletal maturity and the rate of bone loss. This suggests the possibility that the estrogen receptor (ER) gene is one of the candidate genes that determines peak bone density and/or bone turnover rate. We investigated two established restriction fragment length polymorphisms (RFLPs) in intron 1 at the ER gene, represented as PvuII and XbaI. In 598 healthy Korean women aged 20–74 years, we examined the association of these ER genotypes with bone mineral density (BMD) and bone turnover status. The distribution of the PvuII and XbaI RFLPs was as follows: pp 205 (34.3%), Pp 308 (51.5%), PP 85 (14.2%) and xx 384 (64.2%), Xx 180 (30.1%), XX 34 (5.7%), respectively (where capital letters signify the absence of, and lower-case letters signify the presence of, the restriction site of each RFLP). No significant genotypic differences were found in BMD and bone markers. We grouped the subjects into three categories according to their menstrual status: 104 premenopausal women with regular menstruation, 182 perimenopausal women who had amenorrhea of not less than 3 months and not more than 12 months’ duration, and 312 postmenopausal women whose last menstruation was at least 12 months previously. No significant genotypic difference in either BMD or bone markers was found in any of these three groups. Furthermore we categorized women in peri- and postmenopause into a high loser group and a normal loser group according to the level of bone resorption markers. There was no difference in genotypic proportions between the high and normal loser groups. Our data suggest that these ER polymorphisms are not associated with BMD or bone turnover in Korean women. Received: 16 March 1998 / Accepted: 17 August 1998     

Molecularly Defined Lactose Malabsorption,Peak Bone Mass and Bone Turnover Rate in Young Finnish Men

Lactose malabsorption (LM; adult-type hypolactasia), an autosomal recessive condition, results from the down-regulation of the activity of lactase enzyme in the intestinal wall. In previous studies the effect of LM on bone mass, bone turnover rate, development of osteoporosis and osteoporotic fractures has remained controversial. We have recently identified a single nucleotide polymorphism (SNP), a C to T change residing 13910 base pairs upstream of the lactase (LCT) gene at chromosome 2q21-22, which shows complete association with lactase persistence, with the C/C_–13910 genotype defining LM and the genotypes C/T_–13910 and T/T_–13910 lactase persistence. The present study was undertaken to examine the relationship of the C/T_–13910 polymorphism to peak bone mass, bone turnover rate, and stress fractures among young Finnish men. The study population comprised 234 young men, aged 18.3 to 20.6 years, 184 men were recruits of the Finnish Army, and 50 were men of similar age who had postponed their military service for reasons not related to health. Bone mineral content (BMC), density (BMD), and scan area were measured in the lumbar spine and upper femur by dual-energy X-ray absorptiometry (DXA). Blood was sampled for genotyping of the C/T_–13910 polymorphism and determination of serum 25-hydroxyvitamin D (25OHD), intact parathyroid hormone (iPTH), type I procollagen aminoterminal propeptide (PINP), and tartrate-resistant acid phosphatase 5b (TRACP5b). Second-void urine samples were collected for the determination of type I collagen aminoterminal telopeptide (NTX). The prevalence of the C/C_–13910-genotype of these young adults did not differ significantly from the corresponding population prevalence of C/C_–13910 (17.1% vs 18.1%) among Finnish blood donors. Fifteen recruits of the army experienced a stress fracture; 3 of them (20%) had the C/C_–13910-genotype. Calcium intake was similar for the three genotypes as were the unadjusted BMCs, scan areas, and BMDs at different measurement sites. The adjustments for age, height, weight, smoking, alcohol consumption, and physical exercise in the multiple regression analysis did not reveal any significant relationships between the lactase genotypes and BMDs at lumbar (P = 0.16), femoral neck (P = 0.99) or total hip (P = 0.96) sites. Serum 25OHD, iPTH, and bone marker levels were similar for the C/C_–13910 C/T_–13910 and T/T_–13910 genotypes. In summary, in young Finnish men, molecularly defined lactose malabsorption does not alter bone turnover rate and impair the acquisition of peak bone mass. Moreover, the C/C_–13910 genotype does not seem to be a risk factor for stress fractures in army recruits.N. Enattah and V.-V. Välimäki equally contributed to the study.     

Disruption of Four-and-a-Half LIM 2 Decreases Bone Mineral Content and Bone Mineral Density in Femur and Tibia Bones of Female Mice

Four-and-a-half LIM 2 (FHL2) is a member of a family of LIM domain proteins which mediate protein-protein interactions. FHL2 acts as a coactivator and binds to important regulators of bone formation such as insulin-like growth factor binding protein (IGFBP)-5, androgen receptor, and β-catenin. We hypothesized that FHL2 is an important regulator of bone formation. We evaluated growth and skeletal parameters in FHL2 knockout (KO) and wild-type (WT) mice at 4, 8, and 12 weeks of age. At 4 weeks of age, lack of FHL2 reduced femur, tibia, and total bone mineral content (BMC) and body weight in all mice. A gender-by-treatment interaction (P ≤ 0.05) was observed for several parameters due to a greater reduction in females. Specifically, femur BMC was reduced 11–27% at 8 and 12 weeks of age and BMD was reduced 7–13% at all ages in female KO mice (P < 0.05). A similar reduction was observed in the tibias at 8 weeks of age. A 6% reduction (P = 0.07) in femur cortical thickness was observed at 12 weeks of age in female KO mice. Interestingly, a gender-specific reduction in IGFBP-5 expression was observed in the femurs of female KO mice. During differentiation of bone marrow stromal cells into osteoblasts, expression of osteocalcin, alkaline phosphatase, and bone sialoprotein was reduced 47–96% in FHL2 KO cells (P < 0.001). In conclusion, FHL2 is an important regulator of peak bone mass, lack of FHL2 produces gender- and site-specific effects on bone accretion and IGFBP-5 expression, and FHL2 is important for optimal osteoblast differentiation in vitro. The information contained in this publication does not necessarily reflect the position or the policy of the government, and no official endorsement should be inferred. All work was performed in facilities provided by the Department of Veterans Affairs.     

RANKL Blockade Reduces Cachexia and Bone Loss Induced by Non-Metastatic Ovarian Cancer in Mice

Tumor- and bone-derived soluble factors have been proposed to participate in the alterations of skeletal muscle size and function in cachexia. We previously showed that mice bearing ovarian cancer (OvCa) exhibit cachexia associated with marked bone loss, whereas bone-targeting agents, such as bisphosphonates, are able to preserve muscle mass in animals exposed to anticancer drugs. De-identified CT images and plasma samples from female patients affected with OvCa were used for body composition assessment and quantification of circulating cross-linked C-telopeptide type I (CTX-I) and receptor activator of NF-kB ligand (RANKL), respectively. Female mice bearing ES-2 tumors were used to characterize cancer- and RANKL-associated effects on muscle and bone. Murine C2C12 and human HSMM myotube cultures were used to determine the OvCa- and RANKL-dependent effects on myofiber size. To the extent of isolating new regulators of bone and muscle in cachexia, here we demonstrate that subjects affected with OvCa display evidence of cachexia and increased bone turnover. Similarly, mice carrying OvCa present high RANKL levels. By using in vitro and in vivo experimental models, we found that elevated circulating RANKL is sufficient to cause skeletal muscle atrophy and bone resorption, whereas bone preservation by means of antiresorptive and anti-RANKL treatments concurrently benefit muscle mass and function in cancer cachexia. Altogether, our data contribute to identifying RANKL as a novel therapeutic target for the treatment of musculoskeletal complications associated with RANKL-expressing non-metastatic cancers. © 2021 American Society for Bone and Mineral Research (ASBMR).     

Relationships Between the Changes of Serum Levels of OPG and RANKL with Age, Menopause, Bone Biochemical Markers and Bone Mineral Density in Chinese Women Aged 20-75

The correlations between the serum levels of OPG, RANKL with age, menopause, bone markers, and bone mineral densities (BMDs) at the lumbar spine and proximal femur were studied in 504 pre- and postmenopausal Chinese women aged 20–75 years. We found that age was positively and negatively correlated with serum concentrations of OPG (r = 0.442, P < 0.001) and RANKL (r = –0.263, P < 0.001), respectively. Compared with premenopausal women, postmenopausal women showed higher serum OPG levels (107.6 ± 3.0 vs 72.0 ± 1.8 pg/ml, P < 0.001), lower serum RANKL concentrations (4.7 ± 0.4 vs. 5.8 ± 0.3 pg/ml, P < 0.001) and RANKL/OPG ratios (0.045 ± 0. 004 vs. 0.099 ± 0.008, P < 0.001). Neither serum levels of OPG nor RANKL or RANKL/OPG ratio correlated with BMDs after adjustment of age and menopause. They also showed no differences among normal, osteopenic and osteoporotic postmenopausal women. Serum levels of OPG were positively correlated with urinary excretion of NTx (r = 0.1453, P = 0.006). Serum levels of RANKL (r = –0.1928, P < 0.001) and RANKL/OPG ratio (r = –0.1303, P = 0.013) were inversely correlated with serum concentrations of OC. In multiple regression analysis, up to 20% variance (R² = 0.106–0.224) of the OPG-RANKL system in peripheral circulation can be explained by age, menopause and bone markers.These results suggest that although serum OPG and RANKL concentrations were unrelated with BMDs, the age– and menopause– dependent changes of serum OPG and RANKL might be a protective mechanism against the accelerated bone loss in postmenopausal women.     

卵巢切除对鼠及异常应力鼠骨密度影响的比较研究

目的：探讨卵巢切除雌激素缺乏时鼠及异常应力鼠不同部位骨密度的变化规律。材料与方法：３个月龄ＳＤ雌性大鼠６４只，双侧卵巢切除（ＯＶＸ）及行假手术（Ｓｈａｍ－Ｏ）；于术后第１、３、５、７个月各处死８只动物，切取Ｌ１～Ｌ２椎体节段、股骨及胫骨，进行ＢＭＤ（骨矿密度）检测；新生２４～７２小时雌性ＳＤ鼠４８只，建立双后肢大鼠模型，椎体、股骨及胫骨均承受较高的应力。３月龄时再同法分组，术后第２、５、７个月时分别处死动物取上述部位进行ＢＭＤ检测。结果：ＯＶＸ后第１个月椎体ＢＭＤ明显骨丢失（Ｐ＜０．０１），股骨丢失较为缓慢，７个月时方呈现差异（Ｐ＜０．０５）。松质骨区丢失时间提前（Ｐ＜０．０５）。胫骨１个月ＢＭＤ即有差异性（Ｐ＜０．０５），松质骨区则呈持续丢失。结论：鼠ＯＶＸ后ＢＭＤ的丢失速度为腰椎＞胫骨＞股骨；腰椎体＞胫骨上段＞股骨上段。双后肢大鼠组亦呈现同样变化，但腰椎、股骨及胫骨上的应力增加，ＢＭＤ的丢失较缓慢。     

The New Selective Estrogen Receptor Modulator MDL 103,323 Increases Bone Mineral Density and Bone Strength in Adult Ovariectomized Rats

Selective estrogen receptor modulators (SERMs) can prevent the bone loss induced by ovariectomy (OVX), but it is not established whether they can increase bone mass and strength in a curative protocol in ovariectomized osteopenic animals. We investigated the influence of a SERM of the new generation, MDL 103,323, on areal bone mineral density (BMD), as measured by dual-energy X-ray absorptiometry, bone strength and remodeling in OVX osteopenic rats. Nine weeks after OVX, 8-month-old rats were divided into six groups of 10 animals. MDL 103,323 was given by gavage at doses of 0.01, 0.1 or 0.6 mg/kg body weight, 5 days a week. The effect of MDL 103,323 was compared with that of the bisphosphonate pamidronate (APD), which was injected subcutaneously at a dose of 1.6 mmol/kg body weight for 5 days every 4 weeks. Lumbar spine (LS), femoral neck (FN), proximal tibia (PT) and midshaft tibia (MT) BMD, bone strength, and proximal tibia histomorphometry, serum osteocalcin, urinary total deoxypyridinoline and serum insulin-like growth factor I (IGF-I) were measured. After 16 weeks of treatment, BMD changes (means ± SEM) were −11.4 ± 2.2, +4.0 ± 2.1 and +6.4 ± 1.0% respectively in OVX controls, in rats treated with 0.1 mg/kg MDL 103,323 (p<0.05) and in APD-treated rats (p<0.02) at the level of LS; −0.4 ± 1.1, +6.7 ± 1.4, +7.2 ± 1.8% (p<0.01 and NS) at the level of FN; and −2.6 ± 1.2%, +5.8 ± 1.2, +6.9 ± 1.4% (p<0.03 and 0.01) at the level of PT. MDL 103,323-treated animals had a higher trabecular bone volume, a higher number of trabeculae and smaller intertrabecular spaces compared with OVX controls. Vertebral body ultimate strength was 186 ± 13, 292 ± 16, 249 ± 23 N (p<0.05) in OVX controls, MDL 103,323-treated rats and APD-treated rats, respectively. The administration of 0.6 mg/kg of MDL 103,323 did not further increase BMD or bone strength, indicating a bell-shaped dose–response curve. MDL 103,323 lowered plasma osteocalcin concentration and urinary deoxypyridinoline excretion. In rats treated with 0.1 mg/kg MDL 103,323, plasma IGF-I was increased as compared with OVX controls (664 ± 36 ng/ml vs 527 ± 39 ng/ml, p<0.05). In conclusion, these results indicate that this new SERM positively influences BMD and lumbar spine bone strength in estrogen-deficient rats. Received: 30 October 1998 / Accepted: 12 April 1999     

Comparison of Microcomputed Tomographic and Microradiographic Measurements of Cortical Bone Porosity

Cortical bone is perforated by a network of canals that have a significant impact upon its material properties. Microcomputed tomography offers the possibility of noninvasively visualizing and quantifying cortical pores in both two and three dimensions. Establishing how two-dimensional (2D) microcomputed tomographic (CT) analysis compares with conventional methods for analyzing cortical porosity is an important prerequisite for the wider adoption of this technique and the development of three-dimensional (3D) analysis. Therefore, we compared porosity-related parameters from 2D microcomputed tomographic images with those from matching microradiographic sections. Samples from five human femora were scanned at a 10-m resolution and then sequentially sectioned and microradiographed. An average of eight image pairs were produced from each femur (total, n = 41). The repeatability and comparability of the two techniques was assessed for three parameters; cortical porosity (%), mean pore area (m²), and pore density (pores/mm²). For repeatability, no significant difference (P > 0.05) was found between the two methods for cortical porosity and mean pore area; however, pore density differed significantly (P < 0.001). For comparability, the bias (± error) between the methods was found to be 0.51% (±0.31%) for cortical porosity and –155 m² (±293 m²) for mean pore area. The bias for pore density was dependent upon measurement size with microcomputed tomographic images having 14% (±9.3%) fewer pores per millimeter squared. The qualitative and quantitative similarities between the two techniques demonstrated the utility of 2D microcomputed tomographic for cortical porosity analysis. However, the relatively poor results for pore density revealed that a higher resolution (<10 m) is needed to consistently visualize all cortical pores in human bone.     

Bone Turnover in Bone Biopsies of Patients with Low-Energy Cortical Fractures Receiving Bisphosphonates: A Case Series

Recent reports of long-term bisphosphonate-treated patients developing cortical fractures have raised concerns that such fractures may relate to excessive suppression of bone turnover after prolonged use of these drugs. To evaluate the bone histology of patients presenting with cortical fractures after bisphosphonate therapy, we conducted a retrospective analysis of patients treated at Washington University Bone Health Program presenting with a history of low-energy cortical fractures (femoral shaft, pelvis, rib, metatarsal, and ankle), who had received bisphosphonates for at least two consecutive years and had undergone bone biopsy. Fifteen of 54 patients who underwent bone biopsy between November 2004 and March 2007 met the criteria. Of these, 10 patients had findings of suppressed trabecular bone remodeling, as demonstrated by lack of double tetracycline labels. There were no significant differences in bone density, clinical features, and biochemical features between those with suppressed turnover and the other five subjects with normal remodeling. However, the low-turnover group had received bisphosphonates (primarily alendronate) for a significantly longer duration (6.5 ± 0.6 vs. 3.9 ± 0.8 years, P = 0.02). Thus, about two-thirds of patients presenting with cortical fractures while on long-term treatment with bisphosphonates had suppressed turnover. Since the prevalence of such histological findings in nonfracture patients remains unknown, the impact of suppressed bone turnover on the development of cortical fractures cannot be determined. Considering the widespread use of bisphosphonates, it appears that the overall risk of cortical fractures is low. However, there may be a subset of as yet unidentified patients who could be predisposed to this complication.