Astrid Kamilla Stunes

Expression and regulation of resistin in osteoblasts and osteoclasts indicate a role in bone metabolism

The adipose tissue is the site of expression and secretion of a range of biologically active proteins, called adipokines, for example, leptin, adiponectin, and resistin. Leptin has previously been shown to be expressed in osteoblasts and to promote bone mineralization, whereas adiponectin expression is enhanced during osteoblast differentiation. In the present study we explored the possible role of resistin in bone metabolism. We found that resistin is expressed in murine preosteoclasts and preosteoblasts (RAW 264.7, MC3T3‐E1), in primary human bone marrow stem cells and in mature human osteoblasts. The expression of resistin mRNA in RAW 264.7 was increased during differentiation and seemed to be regulated through PKC‐ and PKA‐dependent mechanisms. Recombinant resistin increased the number of differentiated osteoclasts and stimulated NFκB promoter activity, indicating a role in osteoclastogenesis. Resistin also enhanced the proliferation of MC3T3‐E1 cells in a PKA and PKC‐dependent manner, but only weakly interfered with genes known to be upregulated during differentiation of MC3T3‐E1 into osteoblasts. All together, our results indicate that resistin may play a role in bone remodeling. J. Cell. Biochem. 99: 824–834, 2006.

Serotonin and fluoxetine modulate bone cell function in vitro

Recent studies have proposed a role for serotonin and its transporter in regulation of bone cell function. In the present study, we examined the in vitro effects of serotonin and the serotonin transporter inhibitor fluoxetine “Prozac” on osteoblasts and osteoclasts. Human mononuclear cells were differentiated into osteoclasts in the presence of serotonin or fluoxetine. Both compounds affected the total number of differentiated osteoclasts as well as bone resorption in a bell‐shaped manner. RT‐PCR on the human osteoclasts demonstrated several serotonin receptors, the serotonin transporter, and the rate‐limiting enzyme in serotonin synthesis, tryptophan hydroxylase 1 (Tph1). Tph1 expression was also found in murine osteoblasts and osteoclasts, indicating an ability to produce serotonin. In murine pre‐osteoclasts (RAW264.7), serotonin as well as fluoxetine affected proliferation and NFκB activity in a biphasic manner. Proliferation of human mesenchymal stem cells (MSC) and primary osteoblasts (NHO), and 5‐HT2A receptor expression was enhanced by serotonin. Fluoxetine stimulated proliferation of MSC and murine preosteoblasts (MC3T3‐E1) in nM concentrations, µM concentrations were inhibitory. The effect of fluoxetine seemed direct, probably through 5‐HT2 receptors. Serotonin‐induced proliferation of MC3T3‐E1 cells was inhibited by the PKC inhibitor (GF109203) and was also markedly reduced when antagonists of the serotonin receptors 5‐HT2B/C or 5‐HT2A/C were added. Serotonin increased osteoprotegerin (OPG) and decreased receptor activator of NF‐κB ligand (RANKL) secretion from osteoblasts, suggesting a role in osteoblast‐induced inhibition of osteoclast differentiation, whereas fluoxetine had the opposite effect. This study further describes possible mechanisms by which serotonin and the serotonin transporter can affect bone cell function. J. Cell. Biochem. 98: 139–151, 2006.

Adipocytes express a functional system for serotonin synthesis, reuptake and receptor activation

Aims: Serotonergic pathways in the central nervous system (CNS) are activated in the regulation of food intake and body weight. We hypothesized that adipocytes, like other cells of mesenchymal origin, possess serotonin receptors and thus could be regulated by peripherally circulating serotonin.

Different skeletal effects of the peroxisome proliferator activated receptor (PPAR)α agonist fenofibrate and the PPARγ agonist pioglitazone

BackgroundAll the peroxisome proliferator activated receptors (PPARs) are found to be expressed in bone cells. The PPARγ agonist rosiglitazone has been shown to decrease bone mass in mice and thiazolidinediones (TZDs) have recently been found to increase bone loss and fracture risk in humans treated for type 2 diabetes mellitus. The aim of the study was to examine the effect of the PPARα agonist fenofibrate (FENO) and the PPARγ agonist pioglitazone (PIO) on bone in intact female rats.MethodsRats were given methylcellulose (vehicle), fenofibrate or pioglitazone (35 mg/kg body weight/day) by gavage for 4 months. BMC, BMD, and body composition were measured by DXA. Histomorphometry and biomechanical testing of excised femurs were performed. Effects of the compounds on bone cells were studied.ResultsThe FENO group had higher femoral BMD and smaller medullary area at the distal femur; while trabecular bone volume was similar to controls. Whole body BMD, BMC, and trabecular bone volume were lower, while medullary area was increased in PIO rats compared to controls. Ultimate bending moment and energy absorption of the femoral shafts were reduced in the PIO group, while similar to controls in the FENO group. Plasma osteocalcin was higher in the FENO group than in the other groups. FENO stimulated proliferation and differentiation of, and OPG release from, the preosteoblast cell line MC3T3-E1.ConclusionWe show opposite skeletal effects of PPARα and γ agonists in intact female rats. FENO resulted in significantly higher femoral BMD and lower medullary area, while PIO induced bone loss and impairment of the mechanical strength. This represents a novel effect of PPARα activation.

The peroxisome proliferator-activated receptor (PPAR) alpha agonist fenofibrate maintains bone mass, while the PPAR gamma agonist pioglitazone exaggerates bone loss, in ovariectomized rats

BackgroundActivation of peroxisome proliferator-activated receptor (PPAR)gamma is associated with bone loss and increased fracture risk, while PPARalpha activation seems to have positive skeletal effects. To further explore these effects we have examined the effect of the PPARalpha agonists fenofibrate and Wyeth 14643, and the PPARgamma agonist pioglitazone, on bone mineral density (BMD), bone architecture and biomechanical strength in ovariectomized rats.MethodsFifty-five female Sprague-Dawley rats were assigned to five groups. One group was sham-operated and given vehicle (methylcellulose), the other groups were ovariectomized and given vehicle, fenofibrate, Wyeth 14643 and pioglitazone, respectively, daily for four months. Whole body and femoral BMD were measured by dual X-ray absorptiometry (DXA), and biomechanical testing of femurs, and micro-computed tomography (microCT) of the femoral shaft and head, were performed.ResultsWhole body and femoral BMD were significantly higher in sham controls and ovariectomized animals given fenofibrate, compared to ovariectomized controls. Ovariectomized rats given Wyeth 14643, maintained whole body BMD at sham levels, while rats on pioglitazone had lower whole body and femoral BMD, impaired bone quality and less mechanical strength compared to sham and ovariectomized controls. In contrast, cortical volume, trabecular bone volume and thickness, and endocortical volume were maintained at sham levels in rats given fenofibrate.ConclusionsThe PPARalpha agonist fenofibrate, and to a lesser extent the PPARaplha agonist Wyeth 14643, maintained BMD and bone architecture at sham levels, while the PPARgamma agonist pioglitazone exaggerated bone loss and negatively affected bone architecture, in ovariectomized rats.

Long-term serotonin effects in the rat are prevented by terguride

Long-term hyperserotoninemia induces heart valve disease in rats, and cases of cardiac valvulopathies have been reported in patients using ergolines, possibly through activation of the 5-hydroxytryptamine(2B) (5HT(2B)) receptor. The ergoline terguride (transdihydrolisuride) is a 5HT(2B/2C) receptor antagonist. Using a rat model, we have investigated whether terguride could prevent serotonin-induced changes in general and heart disease specifically. During 4 months, twelve Sprague-Dawley rats were given daily subcutaneous serotonin injections; twelve rats received a combination of serotonin injections and terguride by gavage, whereas ten rats were untreated controls. Using echocardiography, rats with aortic insufficiency were found in all 3 groups, while pulmonary insufficiency was only found in two rats injected with serotonin alone. Animals given serotonin alone had significantly higher heart weights compared to the controls (p=0.029) and rats given terguride (p=0.034). Rats injected with serotonin alone developed macroscopic skin changes at the injection sites, histologically identified as orthokeratosis and acanthosis. Terguride completely prevented these changes (p=0.0001, p=0.0003). Liver weights were higher in the animals given serotonin alone compared to controls (p=0.014) and terguride treated animals (p=0.009). Stomach weights were higher in animals given serotonin alone compared to rats given terguride (p=0.012). In the mesenchymal cell-line MC3T3-E1, terguride almost completely inhibited serotonin-induced proliferation (p<0.01). Serotonin increases heart, liver and stomach weights, possibly through enhanced proliferation. Terguride inhibits these effects. We propose that terguride may have beneficial effects in the treatment of diseases such as carcinoid syndrome, where serotonin plays an important pathogenic role.

Increased circulating hepatocyte growth factor (HGF): a marker of epithelial ovarian cancer and an indicator of poor prognosis.

OBJECTIVE Hepatocyte growth factor (HGF) has been described to be increased in different cancers. In the present study we wanted to investigate whether HGF in serum can distinguish between benign and malignant ovarian tumors, and whether serum HGF levels can predict the outcome in patients with ovarian carcinomas. METHODS We included 123 consecutive patients appointed for laparotomy due to a pelvic mass. Preoperative levels of serum cancer antigen 125 (CA 125), HGF and HGF activator (HGFA) were quantified with immunological methods. We performed immunohistochemical analyses of HGFα, HGFβ and the receptor c-Met. Five-year survival of patients with advanced disease (stage III and stage IV) was analyzed with the Kaplan-Meier method. RESULTS Sixty patients had ovarian carcinomas, 23 borderline tumors, and 40 benign ovarian tumors. Patients with ovarian carcinomas had significantly higher preoperative HGF and CA 125 serum levels than patients with benign ovarian tumors, and borderline tumors. Patients with borderline tumors had significantly higher CA 125 values than benign cases. A combination of CA 125 and HGF increased the specificity in predicting carcinoma. We observed abundant HGFα, HGFβ and c-Met expressions in all ovarian tumors. Patients with advanced disease and preoperative serum HGF values ≥2SD above reference value had a shorter disease-free survival than patients with advanced disease and serum HGF <2SD above reference value. CONCLUSIONS HGF in serum is an indicator of ovarian carcinoma in women with a pelvic mass, and of a poor prognosis in advanced ovarian cancer.

Maximal Strength Training in Postmenopausal Women With Osteoporosis or Osteopenia

Abstract Mosti, MP, Kaehler, N, Stunes, AK, Hoff, J, and Syversen, U. Maximal strength training in postmenopausal women with osteoporosis or osteopenia. J Strength Cond Res 27(10): 2879–2886, 2013—Current guidelines recommend weight-bearing activities, preferably strength training for improving skeletal health in patients with osteoporosis. What type of strength training that is most beneficial for these patients is not established. Maximal strength training (MST) is known to improve 1-repetition maximum (1RM) and rate of force development (RFD), which are considered as important covariables for skeletal health. Squat exercise MST might serve as an effective intervention for patients with low bone mass. We hypothesized that 12 weeks of squat exercise MST would improve 1RM and RFD in postmenopausal women with osteoporosis or osteopenia and that these changes would coincide with improved bone mineral density (BMD) and bone mineral content (BMC), and serum markers of bone metabolism. The participants were randomized to a training group (TG, n = 10) or control group (CG, n = 11). The TG underwent 12 weeks of supervised squat exercise MST, 3 times a week, with emphasis on rapid initiation of the concentric part of the movement. The CG was encouraged to follow current exercise guidelines. Measurements included 1RM, RFD, BMD, BMC, and serum bone metabolism markers; type 1 collagen amino-terminal propeptide (P1NP) and type 1 collagen C breakdown products (CTX). At posttest, 8 participants remained in each group for statistical analyses. The TG improved the 1RM and RFD by 154 and 52%, respectively. Lumbar spine and femoral neck BMC increased by 2.9 and 4.9%. The ratio of serum P1NP/CTX tended to increase (p = 0.09), indicating stimulation of bone formation. In conclusion, squat exercise MST improved 1RM, RFD, and skeletal properties in postmenopausal women with osteopenia or osteoporosis. The MST can be implemented as a simple and effective training method for patients with reduced bone mass.

MAXIMAL STRENGTH TRAINING IMPROVES BONE MINERAL DENSITY AND NEUROMUSCULAR PERFORMANCE IN YOUNG ADULT WOMEN

Abstract Mosti, MP, Carlsen, T, Aas, E, Hoff, J, Stunes, AK, and Syversen, U. Maximal strength training improves bone mineral density and neuromuscular performance in young adult women. J Strength Cond Res 28(10): 2935–2945, 2014—Exercise guidelines highlight maximizing bone mass early in life as a strategy to prevent osteoporosis. Which intervention is most effective for this purpose remains unclear. This study investigated the musculoskeletal effects of high acceleration, maximal strength training (MST), in young adult women. Thirty healthy women (22 ± 2 years) were randomly assigned to a training group (TG) and a control group (CG). The TG completed 12 weeks of squat MST, executed at 85–90% of maximal strength 1 repetition maximum (1RM), emphasizing progressive loading and high acceleration in the concentric phase. The CG was encouraged to follow the American College of Sports Medicines exercise guidelines for skeletal health. Measurements included bone mineral density (BMD) and body composition by dual-energy X-ray absorptiometry, dynamic and isometric rate of force development (RFD), and squat 1RM. Serum levels of type 1 collagen amino-terminal propeptide (P1NP), type 1 collagen C breakdown products (CTX), and sclerostin were analyzed by immunoassays. In the TG, lumbar spine and total hip BMD increased by 2.2 and 1.0%, whereas serum P1NP increased by 26.2%. Dynamic RFD and 1RM improved by 81.7 and 97.7%, and isometric RFD improved by 38% at 100 milliseconds. These improvements were significantly greater than those observed in the CG. Within the CG, dynamic RFD and 1RM increased by 27.2 and 12.9% while no other significant changes occurred. These findings suggest that squat MST may serve as a simple, time-efficient strategy to optimize peak bone mass in early adulthood.

Decreased bone mineral density and reduced bone quality in H+/K+ATPase beta‐subunit deficient mice

Proton pump inhibitors (PPIs) are widely used against gastroesophageal reflux disease. Recent epidemiological studies suggest that PPI users have an increased risk of fractures, but a causal relationship has been questioned. We have therefore investigated the skeletal phenotype in H+/K+ATPase beta‐subunit knockout (KO) female mice. Skeletal parameters were determined in 6‐ and 20‐month‐old KO mice and in wild‐type controls (WT). Whole body bone mineral density (BMD) and bone mineral content (BMC) were measured by dual energy X‐ray absorptiometry (DXA). Femurs were examined with µCT analyses and break force were examined by a three‐point bending test. Plasma levels of gastrin, RANKL, OPG, osteocalcin, leptin, and PTH were analyzed. KO mice had lower whole body BMC at 6 months (0.53 vs. 0.59 g, P = 0.035) and at 20 months (0.49 vs. 0.74 g, P < 0.01) compared to WT as well as lower BMD at 6 months (0.068 vs. 0.072 g/cm2, P = 0.026) and 20 months (0.067 vs. 0.077 g/cm2, P < 0.01). Mechanical strength was lower in KO mice at the age of 20 months (6.7 vs. 17.9 N, P < 0.01). Cortical thickness at 20 months and trabecular bone volume% at 6 months were significantly reduced in KO mice. Plasma OPG/RANKL ratio and PTH was increased in KO mice compared to controls. H+/K+ATPase beta subunit KO mice had decreased BMC and BMD, reduced cortical thickness and inferior mechanical bone strength. Whereas the mechanism is uncertain, these findings suggest a causal relationship between long‐term PPI use and an increased risk of fractures. J. Cell. Biochem. 113: 141–147, 2012.