[Treatment of emotion regulation problems in people with neurofibromatosis type 1]. / Behandeling van emotieregulatieproblemen bij mensen met neurofibromatose type 1.

Individuals with the genetic disorder neurofibromatosis type 1 (NF1) are typically diagnosed in a medical hospital setting strongly relying on the presence of well-defined physical symptoms such as neurofibromas or pigmentary spots (known as café-au-lait spots). In mental health care settings, however, aside from a few highly specialized centres, the diagnosis and treatment of individuals with NF1 receives little attention, while the need for psychological treatment is increasingly identified, both in clinical practice and in the scientific literature. Occasional referrals of individuals with NF1 to the mental health services are often only targeted at psychological assessment. Subsequent treatment, however, is usually lacking. We describe two individuals with NF1 for whom by means of specialized clinical neuropsychological assessment, participation in a tailored dialectical behavior therapy (DBT) skills training was indicated. We exposit how they were able to develop their skills and how they themselves and their significant others experienced the treatment.

A comparison of UVb compact lamps in enabling cutaneous vitamin D synthesis in growing bearded dragons.

The effect of exposure to different UVb compact lamps on the vitamin D status of growing bearded dragons (Pogona vitticeps) was studied. Forty-two newly hatched bearded dragons (<24 h old) were allocated to six treatment groups (n = 7 per group). Five groups were exposed to different UVb compact lamps for two hours per day, with a control group not exposed to UVb radiation. At 120 days of age, blood samples were obtained and concentrations of 25(OH)D3 , Ca, P and uric acid were determined. In addition, plasma 25(OH)D3 concentration was determined in free-living adult bearded dragons to provide a reference level. Only one treatment resulted in elevated levels of 25(OH)D3 compared to the control group (41.0 ± 12.85 vs. 2.0 ± 0.0 nmol/L). All UVb-exposed groups had low 25(OH)D3 plasma levels compared to earlier studies on captive bearded dragons as well as in comparison with the free-living adult bearded dragons (409 ± 56 nmol/L). Spectral analysis indicated that all treatment lamps emitted UVb wavelengths effective for some cutaneous vitamin D synthesis. None of these lamps, under this regime, appeared to have provided a sufficient UVb dose to enable synthesis of plasma 25(OH)D3 levels similar to those of free-living bearded dragons in their native habitat.

The diet of free-roaming Australian Central Bearded Dragons (Pogona vitticeps).

The central bearded dragon (Pogona vitticeps) is one of the most popular pet lizards. However, little is known regarding their nutrient requirement, or their natural diet. Therefore, the stomach contents of 14 free-roaming P. vitticeps were determined by flushing. These stomach contents were described taxonomically, and analyzed for crude protein content as well as fatty acid content and composition. Most of the dry matter intake was in the form of animal material (61%) stemming from nine arthropod orders. The most abundant were alates of the termite Drepanotermes sp., accounting for 95% of the total number of prey items and more than half of the total dry matter (DM) intake. Plant material contributed 16% of the total DM intake. The diets were high in crude protein (41-50% DM) and the total fatty acid content was 14-27% of the DM intake. The main fatty acid was C18:1n9c (51-56% of total fatty acids), and polyunsaturated fatty acids (n3 and n6) comprised 6-8% of the total fat intake. Our data suggest that P. vitticeps is an opportunistic predator, which exploits the seasonal availability of prey. Based on our data and other studies, a diet consisting of several insect species, supplemented with leafy vegetables, rich in n3 FA's, would best resemble the expected natural diet of P. vitticeps.

1α,25-dihydroxyvitamin D3 stimulates activin A production to fine-tune osteoblast-induced mineralization.

In healthy bones, mineralization has to be tightly controlled to avoid pathological phenotypes. In this study, we investigated interactions between 1α,25(OH)2 D3 (1,25D3) and activin A in the regulation of osteoblast induced mineralization. In human osteoblast cultures, we demonstrated that besides stimulation of mineralization, 1,25D3 also induced activin A, a strong inhibitor of mineralization. Simultaneously, follistatin (FST), the natural antagonist of activin A, was down-regulated by1,25D3. This resulted in an increase in activin A activity during 1,25D3 treatment. We also showed that in 1,25D3-treated osteoblasts, mineralization can be further increased when activin A activity was abrogated by adding exogenous FST. This observation implies that, besides stimulation of mineralization, 1,25D3 also controls activin A-mediated inhibition of mineralization. Besides activin A, 1,25D3 also induces osteocalcin (BGLAP), another inhibitor of mineralization. Warfarin, which has been shown to inactivate osteocalcin, increased 1,25D3-induced mineralization. Interaction between these two systems became evident from the synergistic increase in BGLAP expression upon blocking activin activity in 1,25D3-treated cultures. In conclusion, we demonstrate that 1,25D3 stimulation of mineralization by human osteoblasts is suppressed by concomitant induction of inhibitors of mineralization. Mineralization induction by 1,25D3 may actually be controlled via interplay with activin A and osteocalcin. Finally, this complex regulation of mineralization substantiates the significance of tight control of mineralization to prevent excessive mineralization and consequently reduction in bone quality and strength.

Evidence of vitamin D and interferon-ß cross-talk in human osteoblasts with 1α,25-dihydroxyvitamin D3 being dominant over interferon-ß in stimulating mineralization.

It is well established that 1α-25-dihydroxyvitamin D3 (1,25D3) regulates osteoblast function and stimulates mineralization by human osteoblasts. The aim of this study was to identify processes underlying the 1,25D3 effects on mineralization. We started with gene expression profiling analyses of differentiating human pre-osteoblast treated with 1,25D3. Bioinformatic analyses showed interferon-related and -regulated genes (ISG) to be overrepresented in the set of 1,25D3-regulated genes. 1,25D3 down-regulated ISGs predominantly during the pre-mineralization period. This pointed to an interaction between the vitamin D and IFN signaling cascades in the regulation of osteoblast function. Separately, 1,25D3 enhances while IFNß inhibits mineralization. Treatment of human osteoblasts with 1,25D3 and IFNß showed that 1,25D3 completely overrules the IFNß inhibition of mineralization. This was supported by analyses of extracellular matrix gene expression, showing a dominant effect of 1,25D3 over the inhibitory effect of IFNß. We identified processes shared by IFNß- and 1,25D3-mediated signaling by performing gene expression profiling during early osteoblast differentiation. Bioinformatic analyses revealed that genes being correlated or anti-correlated with interferon-induced protein with tetratricopeptide repeats 1 (IFIT1) were associated with osteoblast proliferation. In conclusion, the current study demonstrates a cross talk between 1,25D3 and IFNß in osteoblast differentiation and bone formation/mineralization. The interaction is complex and depends on the process but importantly, 1,25D3 stimulation of mineralization is dominant over the inhibitory effect of IFNß. These observations are of potential clinical relevance considering the impact of the immune system on bone metabolism in conditions such as rheumatoid arthritis.

IFNß impairs extracellular matrix formation leading to inhibition of mineralization by effects in the early stage of human osteoblast differentiation.

Osteoimmunology is an emerging field of research focused on the interaction of the immune system and bone. In this study we demonstrate that human osteoblasts are sensitive to the immune cytokine interferon (IFN)ß. Osteoblasts respond to IFNß as shown by the induction of several known IFN target genes such as interferon-induced (IFI) proteins (IFIT1, IFI44L), interferon-stimulated gene factor 3 (ISGF3) complex and the induction of IFNß itself. We demonstrated that IFNß has severe inhibitory effects on mineralization of osteoblast-derived extracellular matrix (ECM). Analysis of the timing of the IFNß effects revealed that committed osteoblasts in early stage of differentiation are most sensitive to IFNß inhibition of mineralization. A single IFNß treatment was as effective as multiple treatments. During the progress of differentiation osteoblasts become desensitized for IFNß. This pinpoints to a complex pattern of IFNß sensitivity in osteoblasts. Focusing on early osteoblasts, we showed that IFNß decreased gene expression of ECM-related genes, such as type I Collagen (COL1A1), fibronectin (FN1), fibullin (FBLN1), fibrillin (FBN2), and laminin (LAMA1). Additionally, ECM produced by IFNß-treated osteoblasts contained less collagen protein. IFNß stimulated gene expression of osteopontin (OPN), annexin2 (ANXA2), and hyaluronan synthase 1 (HAS1), which are important factors in the adhesion of hematopoietic stem cells (HSC) in the HSC niche. In conclusion, IFNß directly modifies human osteoblast function by inhibiting ECM synthesis eventually resulting in delayed bone formation and mineralization and induces a HSC niche supporting phenotype. These effects are highly dependent on timing of treatment in the early phase of osteoblast differentiation.

Age-dependent sex differences in calcium and phosphate homeostasis.

BACKGROUND: Sex differences in calcium and phosphate have been observed. We aimed to assess a relation with age. METHODS: We used the laboratory values of serum calcium, phosphate and albumin from three different samples ( 2005, 2010 and 2014 years) using the hospital information system of Erasmus MC, Rotterdam. The samples were divided into three age groups: 1-17, 18-44 and ≥45 years. Sex differences in calcium and phosphate were analyzed using ANCOVA, adjusting for age and serum albumin. Furthermore, sex by age interactions were determined and we analyzed differences between age groups stratified by sex. RESULTS: In all three samples there was a significant sex × age interaction for serum calcium and phosphate, whose levels were significantly higher in women compared to men above 45 years. No sex differences in the younger age groups were found. In men, serum calcium and phosphate levels were highest in the youngest age group compared to age groups of 18-44 and ≥45 years. In women, serum calcium levels were significantly higher in the age group 1-17 and the age group ≥45 years compared to the 18-44 years age group. In women, serum phosphate was different between the three different age groups with highest level in the group 1-17 years and lowest in the group 18-44 years. CONCLUSION: There are age- dependent sex differences in serum calcium and phosphate. Furthermore, we found differences in serum calcium and phosphate between different age groups. Underlying mechanisms for these age- and sex- differences are not yet fully elucidated.

Continuing medical education during pandemic waves of COVID-19: Consensus from medical faculties in Asia, Australia and Europe.

This article was migrated. The article was marked as recommended. Medical faculties have the responsibility to train tomorrow's doctors and in a crisis face the challenge of delivering students into the workforce promptly and safely. Worldwide, medical faculties have faced unprecedented disruptions from viral outbreaks and pandemics including SARS, Ebola, H1N1 and COVID-19 which bring unique challenges. Currently there is worldwide disruption to medical faculties and medical education due to COVID-19. Despite close links with clinical medicine and the known risks of pandemics, many medical faculties have been caught off guard without pandemic planning in place, to deal with an exponential rise in infections and deaths, overwhelmed health services and widespread community risk of transmission. Assessing transmission risk of COVID-19 in teaching, clinical and community attachments and continuing medical education is paramount as medical faculties face subsequent pandemics waves. Consensus statements based on best available evidence and international expertise from medical faculties in Asia, Australia and Europe were developed to help guide the protection of staff and students, priorities on teaching activities and further educational development. Infection prevention, infection control, contact tracing and medical surveillance are detailed to minimise transmission and to enhance safety. Recommendations on teaching activities planning can enhance responsiveness of medical faculties to tackle subsequent waves of COVID-19 infection. A global approach and dialogue are encouraged.

1Alpha,25-(OH)2D3 acts in the early phase of osteoblast differentiation to enhance mineralization via accelerated production of mature matrix vesicles.

1Alpha,25-dihydroxyitamin D(3) (1,25D3) deficiency leads to impaired bone mineralization. We used the human pre-osteoblastic cell line SV-HFO, which forms within 19 days of culture an extracellular matrix that starts to mineralize around day 12, to examine the mechanism by which 1,25D3 regulates osteoblasts and directly stimulates mineralization. Time phase studies showed that 1,25D3 treatment prior to the onset of mineralization, rather than during mineralization led to accelerated and enhanced mineralization. This is supported by the observation of unaltered stimulation by 1,25D3 even when osteoblasts were devitalized just prior to onset of mineralization and after 1,25D3 treatment. Gene Chip expression profiling identified the pre-mineralization and mineralization phase as two strongly distinctive transcriptional periods with only 0.6% overlap of genes regulated by 1,25D3. In neither phase 1,25D3 significantly altered expression of extracellular matrix genes. 1,25D3 significantly accelerated the production of mature matrix vesicles (MVs) in the pre-mineralization. Duration rather than timing determined the extent of the 1,25D3 effect. We propose the concept that besides indirect effects via intestinal calcium uptake 1,25D3 directly accelerates osteoblast-mediated mineralization via increased production of mature MVs in the period prior to mineralization. The accelerated deposition of mature MVs leads to an earlier onset and higher rate of mineralization. These effects are independent of changes in extracellular matrix protein composition. These data on 1,25D3, mineralization, and MV biology add new insights into the role of 1,25D3 in bone metabolism and emphasize the importance of MVs in bone and maintaining bone health and strength by optimal mineralization status.

Synergistic induction of local glucocorticoid generation by inflammatory cytokines and glucocorticoids: implications for inflammation associated bone loss.

OBJECTIVES: Synovial fibroblasts and osteoblasts generate active glucocorticoids by means of the 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) enzyme. This activity increases in response to proinflammatory cytokines or glucocorticoids. During inflammatory arthritis synovium and bone are exposed to both these factors. This study hypothesised that glucocorticoids magnify the effects of inflammatory cytokines on local glucocorticoid production in both synovium and bone. METHODS: The effects of inflammatory cytokines (IL-1beta/tumour necrosis factor alpha; TNFalpha) and glucocorticoids, alone or combined, were assessed on the expression and activity of 11beta-HSD1 in primary synovial fibroblasts, primary human osteoblasts and MG-63 osteosarcoma cells. A range of other target genes and cell types were used to examine the specificity of effects. Functional consequences were assessed using IL-6 ELISA. RESULTS: In synovial fibroblasts and osteoblasts, treatment with cytokines or glucocorticoids in isolation induced 11beta-HSD1 expression and activity. However, in combination, 11beta-HSD1 expression, activity and functional consequences were induced synergistically to a level not seen with isolated treatments. This effect was seen in normal skin fibroblasts but not foreskin fibroblasts or adipocytes and was only seen for the 11beta-HSD1 gene. Synergistic induction had functional consequences on IL-6 production. CONCLUSIONS: Combined treatment with inflammatory cytokines and glucocorticoids synergistically induces 11beta-HSD1 expression and activity in synovial fibroblasts and osteoblasts, providing a mechanism by which synovium and bone can interact to enhance anti-inflammatory responses by increasing localised glucocorticoid levels. However, the synergistic induction of 11beta-HSD1 might also cause detrimental glucocorticoid accumulation in bone or surrounding tissues.

Evidence for direct effects of prolactin on human osteoblasts: Inhibition of cell growth and mineralization.

Hyperprolactinemia is one of the risk factor of decrease in bone mass which has been believed to be mediated by hypogonadism. However, the presence of prolactin receptor in human osteosarcoma cell line and primary bone cell culture from mouse calvariae supported the hypothesis of a direct prolactin (PRL) action on bone cells. Therefore, the aim of this study was to investigate the role of PRL and its signal transduction pathway in the regulation of bone metabolism via osteoblast differentiation. Human pre-osteoblasts (SV-HFO) that differentiate in a 3-week period from proliferating pre-osteoblasts (days 2-7) to extracellular matrix producing cells (days 7-14) which is eventually mineralized (days 14-21) were used. Concentration of PRL mimicked a lactating period (100 ng/ml) was used to incubate SV-HFO for 21 days in osteogenic medium. Human prolactin receptor mRNA and protein are expressed in SV-HFO. PRL significantly decreased osteoblast number (DNA content) which was due to a decrease in proliferation. PRL increased osteogenic markers, RUNX2 and ALP in early stage of osteoblast differentiation while decreasing it later suggesting a bi-directional effect. Calcium measurement and Alizarin red staining showed a reduction of mineralization by PRL while having neither an effect on osteoblast activity nor RANKL/OPG mRNA ratio. We also demonstrated that PRL action on mineralization was not via PI-3 kinase pathway. The present study provides evidence of a direct effect of prolactin on osteoblast differentiation and in vitro mineralization.

Clostridium glycolicum isolated from a patient with otogenic brain abscesses.

We describe a case of brain abscesses with gas formation following otitis media, for which the patient treated himself by placing clay in his ear. Several microorganisms, including Clostridium glycolicum, were cultured from material obtained from the patient. This is the first report of an infection in an immunocompetent patient associated with this microorganism.

Development of osteoarthritic features in estrogen receptor knockout mice.

OBJECTIVE: Estrogens are suggested to play a role in the development of osteoarthritis as indicated by the increased prevalence in women after menopause. We studied whether deletion of the estrogen receptor (ER) alpha, beta, or both in female mice results in cartilage damage, osteophytosis, and changes in subchondral bone of skeletally mature animals. METHODS: We studied knee joints of 6-month-old female ERalpha-/-, ERbeta-/-, and (double) ERalpha-/-beta-/- mice and their wild type (wt) littermates. The presence and size of osteophytes and osteoarthritic changes in cartilage were analyzed using histology. Changes in subchondral plate and trabecular bone were studied using micro-CT. RESULTS: In ERalpha-/-beta-/- mice, we observed an increase in number and/or size of osteophytes and thinning of the lateral subchondral plate. However, cartilage damage was not different from wt. In ERalpha-/- or ERbeta-/- mice, no significant differences in cartilage damage score, osteophyte formation, or subchondral plate thickness were found. The bone volume fraction of the epiphyseal trabecular bone was unchanged in ERalpha-/- mice, increased in ERbeta-/- mice, and decreased in ERalpha-/-beta-/- mice. CONCLUSIONS: We conclude that deletion of both ERs leads to increased osteophytosis, but deletion of one or both ERs does not lead to overt cartilage damage in 6-month-old mice.

ADAMTS5-/- mice have less subchondral bone changes after induction of osteoarthritis through surgical instability: implications for a link between cartilage and subchondral bone changes.

OBJECTIVE: Osteoarthritis (OA) is characterized by damaged articular cartilage and changes in subchondral bone. Previous work demonstrated aggrecanase-2 deficient (ADAMTS5-/-) mice to be protected from cartilage damage induced by joint instability. This study analyzed whether this protective effect on cartilage is also reflected in the subchondral bone structure. METHODS: Right knee joints from 10-week old male wild type (WT) and ADAMTS5-/- mice received transection of the medial meniscotibial ligament to induce OA, whereas left knees were left unoperated. After 8 weeks knee joints were scanned by micro-CT. The proximal tibia was selected for further analysis. Histology was performed to evaluate cartilage damage and osteoclast presence. RESULTS: ADAMTS5-/- joints had a significantly thinner subchondral plate and less epiphyseal trabecular bone compared to WT joints. Histology confirmed previous findings that ADAMTS5-/- mice have significantly less cartilage damage than WT in the instability-induced OA model. Although the subchondral bone plate became significantly thicker at the medial tibial plateau in operated joints of both genotypes, the percentage increase was significantly smaller in ADAMTS5-/- mice (WT: 20.7+/-4.7%, ADAMTS5-/-: 8.3+/-1.2% compared to the left unoperated control joint). In ADAMTS5-/- animals a significant decrease was found in both Oc.N./BS and Oc.S./BS. Finally, in WT but not in ADAMTS5-/- mice a significant correlation was found between medial subchondral bone plate thickness and cartilage damage at the medial tibial plateau. CONCLUSION: ADAMTS5-/- joints that were protected from cartilage damage showed minor changes in the subchondral bone structure, in contrast to WT mice where substantial changes were found. This finding suggests links between the process of cartilage damage and subchondral bone changes in instability-induced OA.

Wnt signaling acts and is regulated in a human osteoblast differentiation dependent manner.

The Wnt signaling pathway is an important regulator of cellular differentiation in a variety of cell types including osteoblasts. In this study, we investigated the impact of Wnt signaling on the function of human osteoblasts in relation to the stage of differentiation. Differentiating osteoblasts were created upon glucocorticoid (GC) treatment, whereas nondifferentiating osteoblasts were created by excluding GCs from the culture medium. GC-induced differentiation suppressed endogenous beta-catenin levels and transcriptional activity. During GC-induced osteoblast differentiation, activation of Wnt signaling slightly decreased alkaline phosphatase activity, but strongly suppressed matrix mineralization. In addition, mRNA expression of several Wnt signaling related genes was strongly regulated during GC-induced osteoblast differentiation, including frizzled homolog 8, dickkopf homolog 1, and secreted frizzled-related protein 1. In contrast, in the absence of GC-induced differentiation, Wnt signaling acted positively by stimulating basal alkaline phosphatase activity. Interestingly, pre-stimulation of Wnt signaling in early osteoblasts enhanced their differentiation capacity later on during the GC-induced differentiation process. In conclusion, we showed a differentiation-dependent effect of Wnt signaling on osteoblasts. Wnt signaling stimulated early osteoblasts in their capacity to differentiate, whereas mature osteoblasts were strongly inhibited in their capacity to induce mineralization. Moreover, osteoblast differentiation suppressed endogenous Wnt signaling and changed the expression of multiple Wnt signaling related genes.

Subclinical hypocalcaemia in captive Asian elephants (Elephas maximus).

The hypothesis that hypocalcaemia may play a role in dystocia in captive Asian elephants (Elephas maximus) was investigated. The objectives of the study were to measure the total calcium concentration in elephant plasma; assess the changes in parameters of calcium metabolism during a feeding trial; investigate a possible relationship between calcium metabolism and dystocia; and assess bone mineralisation in captive Asian elephants in vivo. The following parameters were measured: total and ionised calcium, inorganic phosphorous and magnesium, the fractional excretions of these minerals, intact parathyroid hormone, 25-OH-D(3) and 1,25-OH-D(3). Radiographs were taken from tail vertebrae for assessment of bone mineralisation. The mean (sd) heparinised plasma total calcium concentration was 2.7 (0.33) mmol/l (n=43) ranging from 0.84 to 3.08 mmol/l in 11 Asian elephants. There was no significant correlation between plasma total calcium concentration and age. Following feeding of a calcium rich ration to four captive Asian elephant cows, plasma total and ionised calcium peaked at 3.6 (0.24) mmol/l (range 3.4 to 3.9 mmol/l) and 1.25 (0.07) mmol/l (range 1.17 to 1.32 mmol/l), respectively. Plasma ionised calcium concentrations around parturition in four Asian elephant cows ranged from 0.37 to 1.1 mmol/l only. The present study indicates that captive Asian elephants might be hypocalcaemic, and that, in captive Asian elephants, the normal plasma concentration of total calcium should actually be around 3.6 mmol/l and normal plasma concentration of ionised calcium around 1.25 mmol/l. Given the fact that elephants absorb dietary calcium mainly from the intestine, it could be concluded that elephants should be fed calcium-rich diets at all times, and particularly around parturition. In addition, normal values for ionised calcium in captive Asian elephants should be reassessed.

Vitamin D receptor genotype is associated with radiographic osteoarthritis at the knee.

Osteoporosis and osteoarthritis are age-related disorders of the skeleton with genetic components. Low bone density is a risk factor for osteoporotic fracture while osteoarthritis is associated with increased bone density. The 1,25-dihydroxyvitamin D3 receptor (VDR) gene locus was previously found to be associated with bone density. We therefore studied the relationship between radiographic osteoarthritis at the knee and VDR genotype in a population-based sample (n = 846), using molecular haplotyping of anonymous intragenic DNA polymorphisms. Radiographic osteoarthritis was defined using the Kellgren score, which is based on the assessment of osteophytes and joint space narrowing (JSN). We show that one VDR haplotype allele is significantly overrepresented in individuals with knee osteoarthritis and associated with a 2.27-fold increased relative risk (95% confidence interval 1.46, 3.52). Adjustment for bone density at the femoral neck did not change these results, indicating that the association is not mediated by bone density. The association appeared to be largely explained by the presence of osteophytes rather than JSN. Our results indicate a role of the VDR gene in the pathogenesis of osteophytes while linkage disequilibrium with another nearby gene, i.e., the collagen type IIa1 gene encoding the most abundant protein in cartilage, might contribute to the association.

Evidence for auto/paracrine actions of vitamin D in bone: 1alpha-hydroxylase expression and activity in human bone cells.

Vitamin D is an important regulator of mineral homeostasis and bone metabolism. 1Alpha-hydroxylation of 25-(OH)D3 to form the bioactive vitamin D hormone, 1alpha,25-(OH)2D3, is classically considered to take place in the kidney. However, 1alpha-hydroxylase has been reported at extrarenal sites. Whether bone is a 1alpha,25-(OH)2D3 synthesizing tissue is not univocal. The aim of this study was to investigate an autocrine/paracrine function for 1alpha,25-(OH)2D3 in bone. We show that 1alpha-hydroxylase is expressed in human osteoblasts, as well as the vitamin D binding protein receptors megalin and cubilin. Functional analyses demonstrate that after incubation with the 1alpha-hydroxylase substrate 25-(OH)D3, the osteoblasts can produce sufficient 1alpha,25-(OH)2D3 to modulate osteoblast activity, resulting in induced alkaline phosphatase (ALP) activity, osteocalcin (OC) and CYP24 mRNA expression, and mineralization. The classical renal regulators of 1alpha-hydroxylase, parathyroid hormone, and ambient calcium do not regulate 1alpha-hydroxylase in osteoblasts. In contrast, interleukin (IL)-1beta strongly induces 1alpha-hydroxylase. Besides the bone-forming cells, we demonstrate 1alpha-hydroxylase activity in the bone resorbing cells, the osteoclasts. This is strongly dependent on osteoclast inducer RANKL. This study showing expression, activity, and functionality of 1alpha-hydroxylase unequivocally demonstrates that vitamin D can act in an auto/paracrine manner in bone.

Osteoclastogenic capacity of peripheral blood mononuclear cells is not different between women with and without osteoporosis.

INTRODUCTION: Peripheral Blood Mononuclear Cells (PBMCs) have been extensively used as a culture model to generate osteoclasts in vitro. The aim of this study was to assess the osteoclastogenic potential of PBMCs derived from post-menopausal women with longstanding osteoporosis and compare this with PBMCs from healthy controls. MATERIAL AND METHODS: We selected from the population-based Rotterdam Study 82 participants of which 43 were diagnosed with osteoporosis (T-score below -2.5 at the lumbar spine) and the presence of at least 1 fracture and 29 healthy controls (T-score above 1; no fracture). PBMCs were differentiated into osteoclasts, and both differentiation capacity and activity were measured. Total RNA was obtained to assess gene expression of osteoclast markers. Deoxypyridinoline (DPD) was measured in plasma as a marker for bone resorption, in vivo. RESULTS: Neither the number of osteoclasts nor cathepsin K (CTSK) and dendritic cell-specific transmembrane protein (TM7SF4) gene expression was significantly different between both groups. There was also no significant difference in resorption pit area and plasma DPD levels. Stratification by fracture type into a group with vertebral, non-vertebral and both vertebral and non-vertebral fractures showed no difference in osteoclast formation or osteoclastic bone resorption. However, plasma DPD, but not the RNA expression markers, was significantly lower in the group of subjects with vertebral fracture group and those with vertebral and non-vertebral fractures compared to the healthy controls. No differences in osteoclastogenesis, osteoclastic resorption and plasma DPD levels were detected also after exclusion of past or present users of bisphosphonates and glucocorticoids. Stratification into high and low DPD levels showed higher osteoclastogenesis and more osteoclastic bone resorption in the high DPD group compared to the low DPD levels within the group of osteoporotic subjects. CONCLUSION: This study showed no difference in PBMC osteoclastogenic capacity and activity between women with and without osteoporosis and at least one previous fracture, who were on average 29.5years after menopause, suggesting that there is no difference in circulating osteoclast precursors. Although we cannot exclude that circulating precursors may behave differently at the bone site, it is possible that long after menopause a more stable phase of bone turnover is reached compared to earlier after the start of menopause in which differences in circulating osteoclast precursors and osteoclastogenic potential are more prominent.

Ghrelin and unacylated ghrelin stimulate human osteoblast growth via mitogen-activated protein kinase (MAPK)/phosphoinositide 3-kinase (PI3K) pathways in the absence of GHS-R1a.

Recent studies demonstrate widespread expression of ghrelin among tissues and have uncovered its pleiotropic nature. We have examined gene expression of ghrelin and its two receptor splice variants, growth hormone secretagogue receptors (GHS-R) 1a and 1b, in human bone biopsies and in the human pre-osteoblastic SV-HFO cell line during differentiation. Additionally, we examined proliferative effects of ghrelin and unacylated ghrelin (UAG) in differentiating and non-differentiating cells. We detected GHS-R1b mRNA in human bone and osteoblasts but not ghrelin's cognate receptor GHS-R1a, using two different real-time PCR assays and both total RNA and mRNA. In osteoblasts GHS-R1b mRNA expression remained low during the first 14 days of culture, but increased 300% in differentiating cells by day 21. Both human bone biopsies and osteoblasts expressed ghrelin mRNA, and osteoblasts were found to secrete ghrelin. Overall, ghrelin gene expression was greater in differentiating than non-differentiating osteoblasts, but was not increased during culture in either group. Ghrelin and UAG induced thymidine uptake dose-dependently, peaking at 1 and 10 nM respectively, at day 6 of culture in both non-differentiating and differentiating osteoblasts. The proliferative response to ghrelin and UAG declined with culture time and state of differentiation. The proliferative effects of ghrelin and UAG were suppressed by inhibitors of extracellular-signal-regulated kinase (ERK) and phosphoinositide-3 kinase, and both peptides rapidly induced ERK phosphorylation. Overall, our data suggest new roles for ghrelin and UAG in modulating human osteoblast proliferation via a novel signal transduction pathway.