Age-associated Iron Accumulation in Bone: Implications for Postmenopausal Osteoporosis and a New Target for Prevention and Treatment by Chelation

Iron accumulation in tissues is believed to be a characteristic of aged humans and a risk factor for some chronic diseases. However, it is not known whether age-associated iron accumulation is part of the pathogenesis of postmenopausal osteoporosis that affects approximately one out three women worldwide. Here, we confirmed that this accumulation of iron was associated with osteopenia in ovariectomized (OVX) rats (a model of peri- and postmenopausal osteoporosis due to estrogen deficiency). To further investigate whether the increased iron level plays a causal role in the onset of bone loss, we treated OVX rats with an orally active and bone targeted chelator that prevented iron accumulation in their skeletal tissues. The results showed that this treatment mitigated the loss of bone mass and the deterioration of bone micro-architecture. We also found that one possible mechanism of the protective action of iron chelation was to significantly reduce bone resorption. Thus, these findings provide a novel target and a potentially useful therapeutic strategy for the prevention and treatment of postmenopausal osteoporosis and perhaps other age-related diseases.     

Icariin protects against iron overload-induced bone loss via suppressing oxidative stress

Iron overload is common in patients with diseases such as hemoglobinopathies, hereditary hemochromatosis or elderly men and postmenopausal women. This disorder is frequently associated with bone loss and recently has been considered as an independent risk factor for osteoporosis. By excess reactive oxygen species (ROS) production through Fenton reaction, iron could induce osteoblast apoptosis, inhibit osteoblast osteogenic differentiation. Moreover, Iron could also promote osteoclasts differentiation and bone absorption. The goal of the study is to investigate whether icariin could reverse iron overload-induced bone loss in vitro and in vivo. Icariin is the major active ingredient of Herba Epimedii and has antioxidant, antiosteoporosis functions. In the current study, we demonstrated that oral administration of icariin significantly prevented bone loss in iron overloaded mice. Icariin could protect against iron overload-induced mitochondrial membrane potential dysfunction and ROS production, promote osteoblast survival and reverse the reduction of Runx2, alkaline phosphatase, and osteopontin expression induced by iron overload. Icariin also inhibited osteoclasts differentiation and function. Moreover, we also found that icariin remarkably reduced iron accumulation in bone marrow, suggesting that icariin has the ability to regulate systemic iron metabolism in vivo. These results indicated that icariin could be a potential natural resource for developing medicines to prevent or treat iron overload-induced osteoporosis.     

Regulation of Osteoblast Differentiation and Iron Content in MC3T3-E1 Cells by Static Magnetic Field with Different Intensities

Many studies have indicated that static magnetic fields (SMFs) have positive effects on bone tissue, including bone formation and bone healing process. Evaluating the effects of SMFs on bone cell (especially osteoblast) function and exploring the mechanism, which is critical for understanding the possible risks or benefits from SMFs to the balance of bone remodeling. Iron and magnetic fields have the natural relationship, and iron is an essential element for normal bone metabolism. Iron overload or deficiency can cause severe bone disorders including osteoporosis. However, there are few reports regarding the role of iron in the regulation of bone formation under SMFs. In this study, hypomagnetic field (HyMF) of 500 nT, moderate SMF (MMF) of 0.2 T, and high SMF (HiMF) of 16 T were used to investigate how osteoblast (MC3T3-E1) responses to SMFs and iron metabolism of osteoblast under SMFs. The results showed that SMFs did not pose severe toxic effects on osteoblast growth. During cell proliferation, iron content of osteoblast MC3T3-E1 cells was decreased in HyMF, but was increased in MMF and HiMF after exposure for 48 h. Compared to untreated control (i.e., geomagnetic field, GMF), HyMF and MMF exerted deleterious effects on osteoblast differentiation by simultaneously retarding alkaline phosphatase (ALP) activity, mineralization and calcium deposition. However, when exposed to HiMF of 16 T, the differentiation potential showed the opposite tendency with enhanced mineralization. Iron level was increased in HyMF, constant in MMF and decreased in HiMF during cell differentiation. In addition, the mRNA expression of transferrin receptor 1 (TFR1) was promoted by HyMF but was inhibited by HiMF. At the same time, HiMF of 16 T and MMF of 0.2 T increased the expression of ferroportin 1 (FPN1). In conclusion, these results indicated that osteoblast differentiation can be regulated by altering the strength of the SMF, and iron is possibly involved in this process.

     

Iron Overload Inhibits Osteoblast Biological Activity Through Oxidative Stress

Iron overload has recently been connected with bone mineral density in osteoporosis. However, to date, the effect of iron overload on osteoblasts remains poorly understood. The purpose of this study is to examine osteoblast biological activity under iron overload. The osteoblast cells (hFOB1.19) were cultured in a medium supplemented with different concentrations (50, 100, and 200 μM) of ferric ammonium citrate as a donor of ferric ion. Intracellular iron was measured with a confocal laser scanning microscope. Reactive oxygen species (ROS) were detected by 2,7-dichlorofluorescin diacetate fluorophotometry. Osteoblast biological activities were evaluated by measuring the activity of alkaline phosphatase (ALP) and mineralization function. Results indicated that iron overload could consequently increase intracellular iron concentration and intracellular ROS levels in a concentration-dependent manner. Additionally, ALP activity was suppressed, and a decline in the number of mineralized nodules was observed in in vitro cultured osteoblast cells. According to these results, it seems that iron overload probably inhibits osteoblast function through higher oxidative stress following increased intracellular iron concentrations.     

Iron overload enhances human mesenchymal stromal cell growth and hampers matrix calcification

BackgroundIron overload syndromes include a wide range of diseases frequently associated with increased morbidity and mortality. Several organs are affected in patients with iron overload including liver, heart, joints, endocrine glands, and pancreas. Moreover, severe bone and hemopoietic tissue alterations are observed. Because of the role of bone marrow mesenchymal stromal cells (BM-MSCs) in bone turnover and hematopoiesis, iron effects on primary BM-MSCs cultures were evaluated.MethodsPrimary human BM-MSCs cultures were prepared and the effects of iron on their proliferation and differentiation were characterized by biochemical analyses and functional approaches.ResultsAddition of iron to the culture medium strongly increased BM-MSCs proliferation and induced their accelerated S phase entry. Iron enters BM-MSCs through both transferrin-dependent and transferrin-independent mechanisms, inducing the accumulation of cyclins E and A, the decrease of p27^Kip1, and the activation of MAPK pathway. Conversely, neither apoptotic signs nor up-regulation of reactive oxygen species were observed. Iron inhibited both differentiation of BM-MSCs into osteoblasts and in vitro matrix calcification. These effects result from the merging of inhibitory activities on BM-MSCs osteoblastic commitment and on the ordered matrix calcification process.ConclusionsWe demonstrated that BM-MSCs are a target of iron overload. Iron accelerates BM-MSCs proliferation and affects BM-MSCs osteoblastic commitment, hampering matrix calcification.General SignificanceOur study reports, for the first time, that iron, at concentration found in overloaded patient sera, stimulates the growth of BM-MSCs, the BM multipotent stromal cell component. Moreover, iron modulates the physiological differentiation of these cells, affecting bone turnover and remodeling.     

Decreased Bone Formation Explains Osteoporosis in a Genetic Mouse Model of Hemochromatosiss

Osteoporosis may complicate iron overload diseases such as genetic hemochromatosis. However, molecular mechanisms involved in the iron-related osteoporosis remains poorly understood. Recent in vitro studies support a role of osteoblast impairment in iron-related osteoporosis. Our aim was to analyse the impact of excess iron in Hfe^-/- mice on osteoblast activity and on bone microarchitecture. We studied the bone formation rate, a dynamic parameter reflecting osteoblast activity, and the bone phenotype of Hfe^−/− male mice, a mouse model of human hemochromatosis, by using histomorphometry. Hfe^−/− animals were sacrificed at 6 months and compared to controls. We found that bone contains excess iron associated with increased hepatic iron concentration in Hfe^−/− mice. We have shown that animals with iron overload have decreased bone formation rate, suggesting a direct impact of iron excess on active osteoblasts number. For bone mass parameters, we showed that iron deposition was associated with bone loss by producing microarchitectural impairment with a decreased tendency in bone trabecular volume and trabecular number. A disorganization of trabecular network was found with marrow spaces increased, which was confirmed by enhanced trabecular separation and star volume of marrow spaces. These microarchitectural changes led to a loss of connectivity and complexity in the trabecular network, which was confirmed by decreased interconnectivity index and increased Minkowski’s fractal dimension. Our results suggest for the first time in a genetic hemochromatosis mouse model, that iron overload decreases bone formation and leads to alterations in bone mass and microarchitecture. These observations support a negative effect of iron on osteoblast recruitment and/or function, which may contribute to iron-related osteoporosis.     

Does high serum iron level induce low bone mass in sickle cell anemia ?

Iron overload is quite common in patients suffering from hemoglobinopathies causing arthropathies, endocrinal affection and neuropathies. Recently low bone mass was added to the list of complications. This study is conducted to find any correlation between serum iron level and low bone mass in sickle cell anemia (SCA). Patients ≥18 years of age with sickle cell anemia, who attended outpatient clinics or admitted to King Fahd University Hospital, Al Khobar, Saudi Arabia,between 1st September 2006 and August 2007 were the subjects of this study. Patients age and sex were documented and body mass index was calculated. Apart from routine hematological tests, serum ferritin, serum Iron level, total estradiol, testosterone level was done. Bone mineral density measurement was done using dual energy X-ray absorptiometry (DEXA) at upper femur and lumbar spine. The data of 100 patients was analyzed, 48 males and 52 females. The mean age was 27.5 ± 6.1 years. In 64 patients (32 males and 32 females) serum iron level was 319.35 μg/dl and the mean serum ferritin level in males and females was within the normal range. Sixty-eight percent of females and 71.8% of males patients in whom serum iron was high had lower bone mass P = < 0.001. Our study shows that SCA patients in whom serum iron level was higher than normal effected bone mass. Further studies are needed to confirm this as a cause of osteoporosis in SCA patients.     

Determinants of bone density in normal women: risk factors for future osteoporosis?

Postmenopausal osteoporosis is an important public health problem in developed countries. Preventive treatment might effect a large reduction in the incidence, but this needs to be applied selectively to those women at increased risk. Loss of bone density results in an increased risk of fractures in the classical sites of vertebrae and proximal femur. A cross sectional study of bone density measurements was carried out in these sites in British women with a modern, precise densitometric technique. Possible predictors and risk factors for bone density were assessed in these women. Bone density was measured by dual photon absorptiometry in 284 apparently healthy women volunteers aged 21 to 68. The values obtained were similar to those obtained from equivalent studies performed in women in the United States. Peak adult bone density had been attained soon after the end of linear skeletal growth. Thereafter there was some decline with age in the proximal femur, but the major fall in bone density in all sites was related to the menopause. Other factors decreasing bone density, and hence increasing risk for osteoporosis, such as low body weight, alcohol and cigarette consumption, nulliparity, lack of previous use of oral contraceptives, and lack of regular exercise, seemed to be important. None, however, could predict satisfactorily women at future risk for osteoporosis. Direct measurements of bone density in the clinically relevant sites are necessary to determine which women should received preventive treatment for postmenopausal osteoporosis. This would help make such treatment more cost effective.     

The role of anorexia nervosa in secondary osteoporosis development with the risk for low energy fractures

Anorexia nervosa (AN) has in recent years become considerably more common. The disease primarily affects girls and young women, also boys and young men. AN is a risk factor for secondary osteoporosis. AN-related metabolic disturbances lead to diminished bone quality and increased risk of fractures. The consequences of low energy fractures are the main causes of death in women with AN. Hormonal disturbances (e.g. hypoestrogenism, increased levels of ghrelin and Y peptide, changes in leptin and endocannabinoid levels), as well as the mechanisms involved in bone resorption (RANK/RANKL/OPG system), are considered to be of great importance for anorectic bone quality. The risk for osteoporotic, non-vertebral fractures in AN patients is significantly higher than in healthy women. Improvement of bone mineral density is possible after substantial body mass increase. Weight loss, in conjunction with a well-balanced, controlled diet, is the key to correct peak bone mass levels, and diminishes the risk for osteoporosis with its consequence of low energy bone fractures.     

In-depth review: is hepcidin a marker for the heart and the kidney?

Iron is an essential trace element involved in oxidation–reduction reactions, oxygen transport and storage, and energy metabolism. Iron in excess can be toxic for cells, since iron produces reactive oxygen species and is important for survival of pathogenic microbes. There is a fine-tuning in the regulation of serum iron levels, determined by intestinal absorption, macrophage iron recycling, and mobilization of hepatocyte stores versus iron utilization, primarily by erythroid cells in the bone marrow. Hepcidin is the major regulatory hormone of systemic iron homeostasis and is upregulated during inflammation. Hepcidin metabolism is altered in chronic kidney disease. Ferroportin is an iron export protein and mediates iron release into the circulation from duodenal enterocytes, splenic reticuloendothelial macrophages, and hepatocytes. Systemic iron homeostasis is controlled by the hepcidin–ferroportin axis at the sites of iron entry into the circulation. Hepcidin binds to ferroportin, induces its internalization and intracellular degradation, and thus inhibits iron absorption from enterocytes, and iron release from macrophages and hepatocytes. Recent data suggest that hepcidin, by slowing or preventing the mobilization of iron from macrophages, may promote atherosclerosis and may be associated with increased cardiovascular disease risk. This article reviews the current data regarding the molecular and cellular pathways of systemic and autocrine hepcidin production and seeks the answer to the question whether changes in hepcidin translate into clinical outcomes of all-cause and cardiovascular mortality, and cardiovascular and renal end-points.

     

4q22.1 Contributes to Bone Mineral Density and Osteoporosis Susceptibility in Postmenopausal Women of Chinese Han Population

Osteoporosis is a multifactorial disease in which genetic determinants are modulated by hormonal, environmental and nutritional factors. An important clinical risk factor in the pathogenesis of osteoporosis is the presence of genetics polymorphism in/around susceptibility genes/regions. This study explored whether the region of 4q22.1, which confers risk of developing osteoporosis in some populations, associated with bone mineral density and osteoporosis susceptibility in postmenopausal women of Han Chinese. We investigated 32 SNPs with minor allele frequencies ≥0.05 between 20 kb upstream and 20 kb downstream (40 kb window) of rs6532023, mapping in the 4q22.1 region, which was reported to be significantly associated with osteoporosis in previous studies. We found that rs6532023 was significantly associated with bone mineral density and osteoporosis (corrected p = 0.015) in our sample, including 440 cases and 640 controls, and allele G was supposed as a risk factor while T worked as a protective factor. Further genotype association analyses suggested a similar pattern (corrected p = 0.040). Additionally, analyses by haplotypes indicated that a haplotype block rs7683315-rs6532023-rs1471400-rs1471403 in the region associated with bone mineral density and osteoporosis (global p = 0.032), and risk haplotype A-G-G-C had almost 1.5-fold increased in the cases. To our knowledge, this is the first report to examine 4q22.1 region polymorphisms and osteoporosis in Han Chinese. Our results provide further evidence for an effect of the region of 4q22.1 on the etiology of osteoporosis and suggest that 4q22.1 may be a genetic risk factor for bone mineral density and osteoporosis.     

Iron absorption in the iron-deficient rat

Iron absorption in the iron-deficient rat was compared with that in the normal rat to better understand the regulation of this dynamic process. It was found that: Iron uptake by the iron-deficient intestinal mucosa was prolonged as a result of slower gastric release, particularly when larger doses of iron were employed. The increased mucosal uptake of ionized iron was not the result of increased adsorption, but instead appeared related to a metabolically active uptake process, whereas the increased mucosal uptake of transferrin iron was associated with increased numbers of mucosal cell membrane transferrin receptors. Mucosal ferritin acted as an iron storage protein, but its iron uptake did not explain the lower iron absorption in the normal rat. Iron loading the mucosal cell (by presenting a large iron dose to the intestinal lumen) decreased absorption for 3 to 4 days. Iron loading of the mucosal cell from circulating plasma transferrin was proportionate to the plasma iron concentration. Mucosal iron content was the composite of iron loading from the lumen and loading from plasma transferrin versus release of iron into the body. These studies imply that an enhanced uptake-throughout mechanism causes the increased iron absorption in the iron-deficient rat. Results were consistent with the existence of a regulating mechanism for iron absorption that responds to change in mucosal cell iron, which is best reflected by mucosal ferritin.     

The role of anorexia nervosa in secondary osteoporosis development with the risk for low energy fractures

Anorexia nervosa (AN) has in recent years become considerably more common. The disease primarily affects girls and young women, and also boys and young men. AN is a risk factor for secondary osteoporosis. AN-related metabolic disturbances lead to diminished bone quality and increased risk of fractures. The consequences of low energy fractures are the main causes of death in women with AN. Hormonal disturbances (e.g. hypoestrogenism, increased levels of ghrelin and Y peptide, changes in leptin and endocannabinoid levels), as well as the mechanisms involved in bone resorption (RANK/RANKL/OPG), are considered to be of great importance for anorectic bone quality. The risk of osteoporotic, non-vertebral fractures in AN patients is significantly higher than in healthy women. An improvement of bone mineral density is possible after substantial body mass increase. Weight loss, in conjunction with a well-balanced, controlled diet, is the key to correct peak bone mass levels, and diminishes the risk of osteoporosis with its consequence of low energy bone fractures. (Pol J Endocrinol 2011; 62 (1): 45-47).     

Osteoporosis,inflammation and ageing

Osteoporosis is a condition characterized by low bone mass and increased bone fragility, putting patients at risk of fractures, which are major causes of morbidity substantially in older people. Osteoporosis is currently attributed to various endocrine, metabolic and mechanical factors. However, emerging clinical and molecular evidence suggests that inflammation also exerts significant influence on bone turnover, inducing osteoporosis. Numerous proinflammatory cytokines have been implicated in the regulation of osteoblasts and osteoclasts, and a shift towards an activated immune profile has been hypothesized as important risk factor. Chronic inflammation and the immune system remodelling characteristic of ageing, as well as of other pathological conditions commonly associated with osteoporosis, may be determinant pathogenetic factors. The present article will review the current perspectives on the interaction between bone and immune system in the elderly, providing an interpretation of osteoporosis in the light of inflamm-ageing.     

Ascorbic acid reduces the frequency of iron induced micronuclei in bone marrow cells of mice

Iron is a potent oxidant that can lead to the formation of genotoxic lipid peroxides. Ascorbic acid, which enhances dietary iron absorption, has been suggested to enhance the oxidant effects of iron and to directly lead to the formation of lipid peroxides. The combined effects of dietary iron and ascorbic acid on genotoxicity were investigated by measuring the frequency of micronuclei in the bone marrow cells of C3H/He mice. In addition, liver iron concentration was measured in all treated groups. Three weeks old mice were fed diets for 3 weeks containing iron at 100 or 300 mg/kg diet in the form of FeSO(4) that were supplemented either with or without ascorbic acid (15 g/kg diet). The results of the bone marrow micronucleus test revealed that the high iron diet resulted in an increased frequency of micronucleated polychromatic erythrocytes (MnPCEs) as compared to low iron. Ascorbic acid supplementation in the low iron diet did not show any effect on incidence of MnPCEs and protected against the increased frequency of MnPCEs induced by the high iron diet. However, liver iron concentration was significantly increased only in the high iron treated and ascorbic acid supplemented group as compared to all other groups. These results demonstrate that ascorbic acid protects against the clastogenic effects of iron.     

Contribution of bone mineral density and bone turnover markers to the estimation of risk of osteoporotic fracture in postmenopausal women

In osteoporosis, the main cause for concern is the increase in the risk of fractures. The level of bone mineral density (BMD) measured by various techniques has been shown to be a strong predictor of fracture risk in postmenopausal women. However, half of patients with incident fractures have BMD value above the diagnostic threshold of osteoporosis defined as a T-score of -2.5 SD or more below the average value of young healthy women. Clearly there is a need for improvement in the identification of patients at risk for fracture. Several prospective studies have shown that an increased bone resorption evaluated by specific biochemical markers was associated with increased risk of the hip, spine and non-vertebral fractures independently of BMD. The use of bone markers in individual patients may be appropriate in some situations, especially in women who are not detected at risk by BMD measurements. For example, in the OFELY study including 668 postmenopausal women followed prospectively over 9 years, we found that among the 115 incident fractures, 54 (47%) actually occurred in non-osteoporotic women. Among these women, the combination of bone markers and history of previous fracture was highly predictive of fracture risk. Thus, bone markers may be used in the assessment of fracture risk in selected cases in which BMD and clinical risk factors are not enough to take a treatment decision. Advances in our knowledge of bone matrix biochemistry, most notably of post-translational modifications in type I collagen, may allow identification of biochemical markers that reflect changes in the material property of bone, which is an important determinant of bone strength. Preliminary in vitro studies indicate that the extent of post-translational modifications of collagen--which can be reflected in vivo by the measurement of the urinary ratio between native and isomerised type I collagen--play a role in determining the mechanical competence of cortical bone, independently of BMD. Further studies in osteoporosis should explore the changes in these biochemical parameters of bone matrix as they may represent a key component of bone quality.     

Disordered hepcidin–ferroportin signaling promotes breast cancer growth

Iron homeostasis is strictly governed in mammals; however, disordered iron metabolism (such as excess iron burden) is recognized as a risk factor for various types of diseases including cancers. Burgeoning evidence indicates that the central signaling of iron homeostasis, the hepcidin–ferroportin axis, is misregulated in cancers. Nonetheless, the mechanisms of misregulated expression of iron-related genes along this signaling in cancers remain largely unknown. In the current study, we found increased levels of serum hepcidin in breast cancer patients. Reduction of hepatic hepcidin through administration of heparin restrained tumorigenic properties of breast tumor cells. Mechanistic investigation revealed that increased iron, bone morphogenetic protein-6 (BMP6) and interleukin-6 (IL-6) jointly promoted the synthesis of hepatic hepcidin. Tumor hepcidin expression was marginally increased in breast tumors relative to adjacent tissues. In contrast, tumor ferroportin concentration was greatly reduced in breast tumors, especially in malignant tumors, compared to adjacent tissues. Elevation of ferroportin concentration inhibited cell proliferation in vitro and in vivo by knocking down tumor hepcidin expression. Additionally, increased IL-6 was demonstrated to jointly enhance the tumorigenic effects of iron through enforcing cell growth. Our combined data overall deciphered the machinery that altered the hepcidin–ferroportin signaling in breast cancers. Thus, targeting the hepcidin–ferroportin signaling would represent a promising therapeutics to restrain breast cancer growth.     

Elemental iron in the anasazi Indians

Excavation of a pithouse village of Anasazi Indians occupied during Pueblo II (A. D. 900-1100) near Littlefield, Arizona, uncovered 14 adult, children and infant burial sites. Iron measurements were made of samples of bone and the sand surrounding the bones at each burial site. Iron in the bones were found to be equivalent to present day normal values. Iron content in the sand surrounding the bones was low and no relationship was noted between the bone iron and the sand iron. These findings are probably valid only for regions where there is good skeletal preservation. The findings support the concept that the diet of these prehistoric Anasazi Indians was nutritionally adequate.