Multiple verification in computational modeling of bone pathologies

We introduce a model checking approach to diagnose the emerging of bone pathologies. The implementation of a new model of bone remodeling in PRISM has led to an interesting characterization of osteoporosis as a defective bone remodeling dynamics with respect to other bone pathologies. Our approach allows to derive three types of model checking-based diagnostic estimators. The first diagnostic measure focuses on the level of bone mineral density, which is currently used in medical practice. In addition, we have introduced a novel diagnostic estimator which uses the full patient clinical record, here simulated using the modeling framework. This estimator detects rapid (months) negative changes in bone mineral density. Independently of the actual bone mineral density, when the decrease occurs rapidly it is important to alarm the patient and monitor him/her more closely to detect insurgence of other bone co-morbidities. A third estimator takes into account the variance of the bone density, which could address the investigation of metabolic syndromes, diabetes and cancer. Our implementation could make use of different logical combinations of these statistical estimators and could incorporate other biomarkers for other systemic co-morbidities (for example diabetes and thalassemia). We are delighted to report that the combination of stochastic modeling with formal methods motivate new diagnostic framework for complex pathologies. In particular our approach takes into consideration important properties of biosystems such as multiscale and self-adaptiveness. The multi-diagnosis could be further expanded, inching towards the complexity of human diseases. Finally, we briefly introduce self-adaptiveness in formal methods which is a key property in the regulative mechanisms of biological systems and well known in other mathematical and engineering areas.Comment: In Proceedings CompMod 2011, arXiv:1109.104

Estrogen Receptor α36 Mediates a Bone-Sparing Effect of 17β-Estrodiol in Postmenopausal Women

Recently, a membrane-based estrogen receptor (ER), ER-α36, was identified and cloned that transduces membrane-initiated estrogen signaling such as activation of the mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) signaling pathway. Here we show that the postmenopausal level of estradiol (E2) induces mitogenic, antiapoptotic, and antiosteogenic effects and proapoptotic effects in postmenopausal osteoblasts and osteoclasts with high levels of ER-α36 expression, respectively. We also found that ER-α36 mediated the effects of postmenopausal-level E2 on proliferation, apoptosis, and differentiation of osteoblasts through transient activation of the MAPK/ERK pathway, whereas ER-α36-mediated postmenopausal-level E2 induces apoptosis of osteoclasts through prolonged activation of the MAPK/ERK pathway with the involvement of reactive oxygen species. We also show that the levels of ER-α36 expression in bone are positively associated with bone mineral density but negatively associated with bone biochemical markers in postmenopausal women. Thus the higher levels of ER-α36 expression are required for preserving bone mass in postmenopausal and menopausal women who become osteoporotic if ER-α36-mediated activities are dysregulated. © 2011 American Society for Bone and Mineral Research

Bone mineral density loss in relation to the final menstrual period in a multiethnic cohort: Results from the Study of Women's Health Across the Nation (SWAN)

The objective of this study was to describe the time of onset and offset of bone mineral density (BMD) loss relative to the date of the final menstrual period (FMP); the rate and amount of BMD decline during the 5 years before and the 5 years after the FMP; and the independent associations between age at FMP, body mass index (BMI), and race/ethnicity with rates of BMD loss during this time interval. The sample included 242 African American, 384 white, 117 Chinese, and 119 Japanese women, pre‐ or early perimenopausal at baseline, who had experienced their FMP and for whom an FMP date could be determined. Loess‐smoothed curves showed that BMD loss began 1 year before the FMP and decelerated (but did not cease) 2 years after the FMP, at both the lumbar spine (LS) and femoral neck (FN) sites. Piecewise, linear, mixed‐effects regression models demonstrated that during the 10‐year observation period, at each bone site, the rates and cumulative amounts of bone loss were greatest from 1 year before through 2 years after the FMP, termed the transmenopause. Postmenopausal loss rates, those occurring between 2 and 5 years after the FMP, were less than those observed during transmenopause. Cumulative, 10‐year LS BMD loss was 10.6%; 7.38% was lost during the transmenopause. Cumulative FN loss was 9.1%; 5.8% was lost during the transmenopause. Greater BMI and African American heritage were related to slower loss rates, whereas the opposite was true of Japanese and Chinese ancestry. © 2012 American Society for Bone and Mineral ResearchPeer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/89478/1/534_ftp.pd

Effects of 25-Hydroxyvitamin D3 on Proliferation and Osteoblast Differentiation of Human Marrow Stromal Cells Require CYP27B1/1α-Hydroxylase

1,25-Dihydroxyvitamin D3 [1,25(OH)2D3] has many noncalcemic actions that rest on inhibition of proliferation and promotion of differentiation in malignant and normal cell types. 1,25(OH)2D3 stimulates osteoblast differentiation of human marrow stromal cells (hMSCs), but little is known about the effects of 25-hydroxyvitamin D3 [25(OH)D3] on these cells. Recent evidence shows that hMSCs participate in vitamin D metabolism and can activate 25(OH)D3 by CYP27B1/1α-hydroxylase. These studies test the hypothesis that antiproliferative and prodifferentiation effects of 25(OH)D3 in hMSCs depend on CYP27B1. We studied hMSCs that constitutively express high (hMSCshi-1α) or low (hMSCslo-1α) levels of CYP27B1 with equivalent expression of CYP24A1 and vitamin D receptor. In hMSCshi-1α, 25(OH)D3 reduced proliferation, downregulated proliferating cell nuclear antigen (PCNA), upregulated p21Waf1/Cip1, and decreased cyclin D1. Unlike 1,25(OH)2D3, the antiapoptotic effects of 25(OH)D3 on Bax and Bcl-2 were blocked by the P450 inhibitor ketoconazole. The antiproliferative effects of 25(OH)D3 in hMSCshi-1α and of 1,25(OH)2D3 in both samples of hMSCs were explained by cell cycle arrest, not by increased apoptosis. Stimulation of osteoblast differentiation in hMSCshi-1α by 25(OH)D3 was prevented by ketoconazole and upon transfection with CYP27B1 siRNA. These data indicate that CYP27B1 is required for 25(OH)D3's action in hMSCs. Three lines of evidence indicate that CYP27B1 is required for the antiproliferative and prodifferentiation effects of 25(OH)D3 on hMSCs: Those effects were not seen (1) in hMSCs with low constitutive expression of CYP27B1, (2) in hMSCs treated with ketoconazole, and (3) in hMSCs in which CYP27B1 expression was silenced. Osteoblast differentiation and skeletal homeostasis may be regulated by autocrine/paracrine actions of 25(OH)D3 in hMSCs. © 2011 American Society for Bone and Mineral Research

Endocortical bone loss in osteoporosis: The role of bone surface availability

Age-related bone loss and postmenopausal osteoporosis are disorders of bone remodelling, in which less bone is reformed than resorbed. Yet, this dysregulation of bone remodelling does not occur equally in all bone regions. Loss of bone is more pronounced near and at the endocortex, leading to cortical wall thinning and medullary cavity expansion, a process sometimes referred to as "trabecularisation" or "cancellisation". Cortical wall thinning is of primary concern in osteoporosis due to the strong deterioration of bone mechanical properties that it is associated with. In this paper, we examine the possibility that the non-uniformity of microscopic bone surface availability could explain the non-uniformity of bone loss in osteoporosis. We use a computational model of bone remodelling in which microscopic bone surface availability influences bone turnover rate and simulate the evolution of the bone volume fraction profile across the midshaft of a long bone. We find that bone loss is accelerated near the endocortical wall where the specific surface is highest. Over time, this leads to a substantial reduction of cortical wall thickness from the endosteum. The associated expansion of the medullary cavity can be made to match experimentally observed cross-sectional data from the Melbourne Femur Collection. Finally, we calculate the redistribution of the mechanical stresses in this evolving bone structure and show that mechanical load becomes critically transferred to the periosteal cortical bone.Comment: 13 pages, 3 figures. V2: minor stylistic improvements in text/figures; more accurately referenced subsection "Internal mechanical stress distribution"; some improved remarks in the Discussion sectio

Influence of Habitual Physical Behavior – Sleeping, Sedentarism, Physical Activity – On Bone Health in Community-Dwelling Older People

Sedentary behavior (SB) has emerged as an independent public-health risk and may contribute to the lower bone mineral density (BMD) in old (>60 years of age) than young adults. The purpose of this study was to quantify SB and habitual physical behavior (PB) in community-dwelling older adults and how this correlates with BMD. In 112 relatively healthy and independent-living individuals aged 72.5 ± 6.4 years, BMD, PB and SB were determined using dual energy X-ray absorptiometry and 7-day three-dimensional accelerometry, respectively. In men, only healthy and osteopenic BMDs were found, whereas in women, osteoporotic BMD classifications also occurred. Our sample spent ∼61%, 7%, 12% and 19% of daily waking hours in SB, standing, LIPA [light intensity physical activity (PA)] and MVPA (medium-to-vigorous intensity PA), respectively. In men, after accounting for covariates (BMI, total fat, android:gynoid ratio), sleeping (hours/day), number of breaks in SB, number of SB ≥ 5 min, number of PA bouts, total duration of PA bouts (min), mean PA bouts duration (min), LIPA (%PA bout time) and MVPA (%PA bout time) were all predictors of BMD. In women, after accounting for covariates (age, BMI, total fat, android:gynoid ratio), SB (hours/day), SB (% waking hours), LIPA (hours/day), LIPA (% waking hours), MVPA (% waking hours) and number of short SB (i.e., <5 min), total time spent in PA (min) significantly correlated with BMD. In conclusion, the PB predictors of bone health in older persons include: night time sleeping duration, number of short bouts of SB, number and duration of bouts of PA relative to total waking hours. While radar graphs of PB patterns for healthy, osteopenic, osteoporotic individuals highlighted significant differences in PB between them, they were not consistent with the expectations from the Mechanostat Theory: i.e., more loading leads to better bone. Rather, our results suggest that a balance of activities must be maintained across the PB spectrum, where certain PB parameters are especially impactful in each sex, supporting the recently coined multifactorial-based variations in the Mechanostat threshold

Reproductive hormone analyses and effects of adjuvant zoledronic acid in early breast cancer – an AZURE (BIG 01/04) sub-study

Purpose: Adjuvant bisphosphonates have been shown to improve disease outcomes in early breast cancer in women who are postmenopausal at the start of treatment. We explored the influence of pretreatment serum levels of reproductive hormones in the hypothalamic-pituitary-gonadal (HPG) axis from a subset of patients included in the AZURE trial to investigate their impact on disease recurrence and whether reproductive hormone measurements are of value in selecting patients for treatment with adjuvant zoledronic acid. Patients and methods: The AZURE trial is an academic, multi-centre, international phase III trial that randomised patients to standard adjuvant therapy (chemotherapy and/or endocrine therapy) +/- intravenous zoledronic acid, 4 mg for 5 years. Serum from 865 patients taken at randomisation was stored at -80°C prior to central batch analysis for inhibin A, oestradiol and follicle stimulating hormone (FSH). We assessed the clinical value of pretreatment hormone levels for predicting invasive disease free survival (IDFS), skeletal recurrence and distant recurrence and response to treatment with zoledronic acid. Results: Oestradiol in the postmenopausal range (26IU/l) was associated with a longer time to bone as first recurrence (HR 0.66 95%CI: 0.41–1.04 p=0.072) compared to an FSH ≤26IU/l. When all 3 hormone levels were within the assay specified postmenopausal range, a trend to improved IDFS was seen with addition of zoledronic acid in biochemically postmenopausal women only (postmenopausal HR=0.81; 95%CI: 0.54–1.22, non-postmenopausal HR=0.99; 95%CI: 0.69–1.39) with risk reductions that mirrored the results of the main AZURE study, although the interaction between menopausal status and treatment effect was not statistically significant (p=0.47). Conclusion: Oestradiol and FSH may influence the pattern of disease recurrence with postmenopausal levels possibly creating a less conducive environment for the formation of bone metastases, therefore disseminated tumour cells could seek alternative niches outside of bone. Biochemical evaluation of a panel of reproductive hormones may be helpful to assist selection of patients for adjuvant zoledronic acid when menopausal status is unknown

Sex-specific autophagy modulation in osteoblastic lineage: a critical function to counteract bone loss in female

Age-related bone loss is associated with an increased oxidative stress which is worsened by estrogen fall during menauposis. This observation has drawn attention to autophagy, a major cellular catabolic process, able to alleviate oxidative stress in osteoblasts (OB) and osteocytes (OST), two key bone cell types. Moreover, an autophagy decline can be associated with aging, suggesting that an age-related autophagy deficiency in OB and/or OST could contribute to skeletal aging and osteoporosis onset. In the present work, autophagy activity was analyzed in OST and OB in male and female mice according to their age and hormonal status. In OST, autophagy decreases with aging in both sexes. In OB, although a 95% decrease in autophagy is observed in OB derived from old females, this activity remains unchanged in males. In addition, while ovariectomy has no effect on OB autophagy levels, orchidectomy appears to stimulate this process. An inverse correlation between autophagy and the oxidative stress level was observed in OB derived from males or females. Finally, using OBspecific autophagy-deficient mice, we showed that autophagy deficiency aggravates the bone loss associated with aging and estrogen deprivation. Taken together, our data indicate that autophagic modulation in bone cells differs according to sex and cell type. The lowering of autophagy in female OB, which is associated with an increased oxidative stress, could play a role in osteoporosis pathophysiology and suggests that autophagy could be a new therapeutic target for osteoporosis in women

LIS1 Regulates Osteoclast Formation and Function through Its Interactions with Dynein/Dynactin and Plekhm1

Microtubule organization and lysosomal secretion are both critical for the activation and function of osteoclasts, highly specialized polykaryons that are responsible for bone resorption and skeletal homeostasis. Here, we have identified a novel interaction between microtubule regulator LIS1 and Plekhm1, a lysosome-associated protein implicated in osteoclast secretion. Decreasing LIS1 expression by shRNA dramatically attenuated osteoclast formation and function, as shown by a decreased number of mature osteoclasts differentiated from bone marrow macrophages, diminished resorption pits formation, and reduced level of CTx-I, a bone resorption marker. The ablated osteoclast formation in LIS1-depleted macrophages was associated with a significant decrease in macrophage proliferation, osteoclast survival and differentiation, which were caused by reduced activation of ERK and AKT by M-CSF, prolonged RANKL-induced JNK activation and declined expression of NFAT-c1, a master transcription factor of osteoclast differentiation. Consistent with its critical role in microtubule organization and dynein function in other cell types, we found that LIS1 binds to and colocalizes with dynein in osteoclasts. Loss of LIS1 led to disorganized microtubules and aberrant dynein function. More importantly, the depletion of LIS1 in osteoclasts inhibited the secretion of Cathepsin K, a crucial lysosomal hydrolase for bone degradation, and reduced the motility of osteoclast precursors. These results indicate that LIS1 is a previously unrecognized regulator of osteoclast formation, microtubule organization, and lysosomal secretion by virtue of its ability to modulate dynein function and Plekhm1

Investigation of bone resorption within a cortical basic multicellular unit using a lattice-based computational model

In this paper we develop a lattice-based computational model focused on bone resorption by osteoclasts in a single cortical basic multicellular unit (BMU). Our model takes into account the interaction of osteoclasts with the bone matrix, the interaction of osteoclasts with each other, the generation of osteoclasts from a growing blood vessel, and the renewal of osteoclast nuclei by cell fusion. All these features are shown to strongly influence the geometrical properties of the developing resorption cavity including its size, shape and progression rate, and are also shown to influence the distribution, resorption pattern and trajectories of individual osteoclasts within the BMU. We demonstrate that for certain parameter combinations, resorption cavity shapes can be recovered from the computational model that closely resemble resorption cavity shapes observed from microCT imaging of human cortical bone.Comment: 17 pages, 11 figures, 1 table. Revised version: paper entirely rewritten for a more biology-oriented readership. Technical points of model description now in Appendix. Addition of two new figures (Fig. 5 and Fig. 9) and removal of former Fig.