Infectious etiologies among post-donation deferrals in a military blood donation center.

BACKGROUND: The burden of transfusion-transmitted infections among blood recipients remains low due to extensive pre- and post-donation screening. However, the military has the unique challenge of providing blood in austere environments with limited testing capabilities. This study evaluates the infectious etiologies of deferred blood donors at a large military blood donation center. METHODS: All blood donors at the Armed Service Blood Bank Center, San Antonio, between 2017 and 2022 with positive post-donation screening for hepatitis C (HCV), hepatitis B (HBV), human immunodeficiency virus (HIV), human T-lymphotropic virus (HTLV-I/II), Zika (2018-2021), West Nile virus, Trypanosoma cruzi, Treponema pallidum, or Babesia microti (2020-2022) were evaluated. Donors were deferred based on Food and Drug Administration (FDA) guidance. RESULTS: Two-hundred and thirteen (213) donors met FDA criteria for deferral. T. pallidum (n = 45, 50.3 per 100,000), HCV (n = 34, 38.0 per 100,000), and HBV (n = 19, 21.2 per 100,000) were the most common pathogens among those with both positive screening and confirmatory testing. The majority of HIV (95%), Chagas (78%), HTLV-I/II (50%) deferrals were due to indeterminate confirmatory tests following initial positive screens. The majority of deferrals for HBV were for a second occurrence of a positive screen despite negative confirmatory testing. CONCLUSION: The rates of post-donation deferral for transfusion-transmissible infections were low in this military cohort. Our findings suggest that donor testing in deployed service members should focus on HBV, HCV, and T. pallidum and highlight the need for better diagnostics for HIV, Chagas, and HTLV-I/II.

Exercise intensity and oxygen uptake kinetics in African-American and Caucasian women.

The effect of exercise intensity on the on- and off-transient kinetics of oxygen uptake (VO(2)) was investigated in African American (AA) and Caucasian (C) women. African American (n = 7) and Caucasian (n = 6) women of similar age, body mass index and weight, performed an incremental test and bouts of square-wave exercise at moderate, heavy and very heavy intensities on a cycle ergometer. Gas exchange threshold (LT(GE)) was lower in AA (13.6 ± 2.3 mL kg(-1) min(-1)) than C (18.6 ± 5.6 mL kg(-1) min(-1)). The dynamic exercise and recovery VO(2) responses were characterized by mathematical models. There were no significant differences in (1) peak oxygen uptake (VO(2peak)) between AA (28.5 ± 5 mL kg(-1) min(-1)) and C (31.1 ± 6.6 mL kg(-1) min(-1)) and (2) VO(2) kinetics at any exercise intensity. At moderate exercise, the on- and off- VO(2) kinetics was described by a monoexponential function with similar time constants τ (1,on) (39.4 ± 12.5; 38.8 ± 15 s) and τ (1,off) (52.7 ± 10.1; 40.7 ± 4.4 s) for AA and C, respectively. At heavy and very heavy exercise, the VO(2) kinetics was described by a double-exponential function. The parameter values for heavy and very heavy exercise in the AA group were, respectively: τ (1,on) (47.0 ± 10.8; 44.3 ± 10 s), τ (2,on) (289 ± 63; 219 ± 90 s), τ (1,off) (45.9 ± 6.2; 50.7 ± 10 s), τ (2,off) (259 ± 120; 243 ± 93 s) while in the C group were, respectively: τ (1,on) (41 ± 12; 43.2 ± 15 s); τ (2, on) (277 ± 81; 215 ± 36 s), τ (1,off) (40.2 ± 3.4; 42.3 ± 7.2 s), τ (2,off) (215 ± 133; 228 ± 64 s). The on- and off-transients were symmetrical with respect to model order and dependent on exercise intensity regardless of race. Despite similar VO(2) kinetics, LT(GE) and gain of the VO(2) on-kinetics at moderate intensity were lower in AA than C. However, generalization to the African American and Caucasian populations is constrained by the small subject numbers.

A double pre-selection method for natural background levels assessment in coastal groundwater bodies.

To evaluate the chemical status of groundwater bodies (GWB) according to the European Groundwater Directive, EU Member States are required to take into account natural background levels (NBLs) where needed. Assessing the NBLs in coastal GWBs is complicated by seawater intrusion which can be amplified by groundwater withdrawals increasing the salinization of such groundwater systems. This paper proposes a new method for the NBLs assessment in coastal areas based on a double pre-selection (PS) with fixed/dynamic limits. A case study in the Apulia region, located in southeastern Italy, is proposed, where we investigated four adjacent GWBs which form the complex karst, fractured Murgia aquifer, hosted in the Jurassic-Cretaceous carbonate platform, bounded by two seas and sustained by saltwater of marine intrusion in the coastal areas. Data related to 139 monitoring stations (MSs) of the regional groundwater monitoring network were used. The first PS, "static", based on a fixed limit of anthropogenic contamination markers (NO3 and NH4), allows for the elimination of MSs impacted by human activities. On these, the second PS, "dynamic", based on the identification of Cl anomalous values, allows for the identification of additional MSs affected by saline contamination. The residual dataset of MSs was used for the definition of NBLs of Cl, SO4, F and B. A statistical comparison with historical Cl observations finally allowed us to verify if the salinity of current groundwater is representative of pristine conditions. The calculated NBLs of salinity parameters are higher for the two coastal GWBs, with chloride values between 0.8 and 2 mg/L. Conversely, fluorides always show very low NBLs. The double PS approach seems more effective for NBLs calculation in coastal aquifers affected by saline contamination, where the use of a fixed Cl limit fails. It may respond to the international needs for a standardized procedure for NBL assessment.

Global CO2 emissions from dry inland waters share common drivers across ecosystems.

Many inland waters exhibit complete or partial desiccation, or have vanished due to global change, exposing sediments to the atmosphere. Yet, data on carbon dioxide (CO2) emissions from these sediments are too scarce to upscale emissions for global estimates or to understand their fundamental drivers. Here, we present the results of a global survey covering 196 dry inland waters across diverse ecosystem types and climate zones. We show that their CO2 emissions share fundamental drivers and constitute a substantial fraction of the carbon cycled by inland waters. CO2 emissions were consistent across ecosystem types and climate zones, with local characteristics explaining much of the variability. Accounting for such emissions increases global estimates of carbon emissions from inland waters by 6% (~0.12 Pg C y-1). Our results indicate that emissions from dry inland waters represent a significant and likely increasing component of the inland waters carbon cycle.

Effect of soy nuts on adhesion molecules and markers of inflammation in hypertensive and normotensive postmenopausal women.

Recently, it was shown that substituting soy nuts for nonsoy protein in a therapeutic lifestyle change (TLC) diet lowered systolic and diastolic blood pressure by 9.9% and 6.8%, respectively, in postmenopausal women with hypertension and by 5.2% and 2.9%, respectively, in normotensive postmenopausal women. In this study, to examine mechanisms for these reductions, markers of inflammation were measured, including soluble vascular cell adhesion molecule-1, soluble intercellular adhesion molecule-1, C-reactive protein, interleukin-6, and matrix metalloproteinase-9. Sixty healthy postmenopausal women (48 normotensive and 12 with hypertension) were randomized in a crossover design to a TLC diet alone or a TLC diet in which 0.5 cups of soy nuts (25 g soy protein and 101 mg aglycone isoflavones) replaced 25 g of nonsoy protein daily. Each diet was followed for 8 weeks. Compared with the TLC diet alone, levels of soluble vascular cell adhesion molecule-1 were significantly lower on the soy diet in women with hypertension (623.6 +/- 153.8 vs 553.8 +/- 114.4 ng/ml, respectively, p = 0.003), whereas no significant differences were observed in normotensive women. Soy nuts were associated with a trend toward reduction in C-reactive protein in normotensive women. No effect on levels of soluble intercellular adhesion molecule-1, interleukin-6, or matrix metalloproteinase-9 was observed. In conclusion, the reduction in soluble vascular cell adhesion molecule-1 with soy nuts in women with hypertension suggests an improvement in endothelial function that may reflect an overall improvement in the underlying inflammatory process underlying atherosclerosis.

Constitutive activity of the osteoblast Ca2+-sensing receptor promotes loss of cancellous bone.

Changes in extracellular [Ca2+] modulate the function of bone cells in vitro via the extracellular Ca2+-sensing receptor (CaR). Within bone microenvironments, resorption increases extracellular [Ca2+] locally. To determine whether enhanced CaR signaling could modulate remodeling and thereby bone mass in vivo, we generated transgenic mice with a constitutively active mutant CaR (Act-CaR) targeted to their mature osteoblasts by the 3.5 kb osteocalcin promoter. Longitudinal microcomputed tomography of cancellous bone revealed reduced bone volume and density, accompanied by a diminished trabecular network, in the Act-CaR mice. The bone loss was secondary to an increased number and activity of osteoclasts, demonstrated by histomorphometry of secondary spongiosa. Histomorphometry, conversely, indicates that bone formation rates were unchanged in the transgenic mice. Constitutive signaling of the CaR in mature osteoblasts resulted in increased expression of RANK-L (receptor activator of nuclear factor-kappaB ligand), the major stimulator of osteoclast differentiation and activation, which is the likely underlying mechanism for the bone loss. The phenotype of Act-CaR mice is not attributable to systemic changes in serum [Ca2+] or PTH levels. We provide the first in vivo evidence that increased signaling by the CaR in mature osteoblasts can enhance bone resorption and further propose that fluctuations in the [Ca2+] within the bone microenvironment may modulate remodeling via the CaR.

Type B gamma-aminobutyric acid receptors modulate the function of the extracellular Ca2+-sensing receptor and cell differentiation in murine growth plate chondrocytes.

Extracellular calcium-sensing receptors (CaRs) and metabotropic or type B gamma-aminobutyric acid receptors (GABA-B-Rs), two closely related members of family C of the G protein-coupled receptor superfamily, dimerize in the formation of signaling and membrane-anchored receptor complexes. We tested whether CaRs and two GABA-B-R subunits (R1 and R2) are expressed in mouse growth plate chondrocytes (GPCs) by PCR and immunocytochemistry and whether interactions between these receptors influence the expression and function of the CaR and extracellular Ca(2+)-mediated cell differentiation. Both CaRs and the GABA-B-R1 and -R2 were expressed in the same zones of the growth plate and extensively colocalized in intracellular compartments and on the membranes of cultured GPCs. The GABA-B-R1 co-immunoprecipitated with the CaR, confirming a physical interaction between the two receptors in GPCs. In vitro knockout of GABA-B-R1 genes, using a Cre-lox recombination strategy, blunted the ability of high extracellular Ca(2+) concentration to activate phospholipase C and ERK1/2, suppressed cell proliferation, and enhanced apoptosis in cultured GPCs. In GPCs, in which the GABA-B-R1 was acutely knocked down, there was reduced expression of early chondrocyte markers, aggrecan and type II collagen, and increased expression of the late differentiation markers, type X collagen and osteopontin. These results support the idea that physical interactions between CaRs and GABA-B-R1s modulate the growth and differentiation of GPCs, potentially by altering the function of CaRs.

Effect of onsite dietitian counseling on weight loss and lipid levels in an outpatient physician office.

We examined the effect of an outpatient office-based diet and exercise counseling program on weight loss and lipid levels with an onsite dietitian who sees patients at the same visit with the physician and is fully reimbursable. Eighty overweight or obese patients (average age 55 +/- 12 years, baseline body mass index 30.1 +/- 6.4 kg/m(2)) with > or =1 cardiovascular risk factor (86%) or coronary heart disease (14%) were counseled to exercise 30 minutes/day and eat a modified Dietary Approaches to Stop Hypertension (DASH) diet (saturated fat <7%, polyunsaturated fat to 10%, monounsaturated fat to 18%, low in glycemic index and sodium and high in fiber, low-fat dairy products, fruits, and vegetables). Weight, body mass index, lipid levels, and blood pressure were measured at 1 concurrent follow-up visit with the dietitian and physician and > or =1 additional follow-up with the physician. Maximum weight lost was an average of 5.6% (10.8 lb) at a mean follow-up of 1.75 years. Sixty-four (81%) of these patients maintained significant weight loss (average weight loss 5.3%) at a mean follow-up of 2.6 years. Average decrease in low-density lipoprotein cholesterol was 9.3%, average decrease in triglycerides was 34%, and average increase in high-density lipoprotein cholesterol was 9.6%. Systolic blood pressure was lowered from 129 to 126 mm Hg (p = 0.21) and diastolic blood pressure from 79 to 75 mm Hg (p = 0.003). In conclusion, having a dietitian counsel patients concurrently with a physician in the outpatient setting is effective in achieving and maintaining weight loss and is fully reimbursable.

The association between soy nut consumption and decreased menopausal symptoms.

BACKGROUND: Epidemiological studies suggest a low incidence of hot flashes in populations that consume dietary soy. The present study examined the effect of soy nuts on hot flashes and menopausal symptoms. METHODS: Sixty healthy postmenopausal women were randomized in a crossover design to a therapeutic lifestyle changes (TLC) diet alone and a TLC diet of similar energy, fat, and protein content in which one-half cup soy nuts divided into three or four portions spaced throughout the day (containing 25 g soy protein and 101 mg aglycone isoflavones) replaced 25 g of nonsoy protein. During each 8-week diet period, subjects recorded the number of hot flashes and amount of exercise daily. At the end of each 8-week diet period, subjects filled out the menopausal symptom quality of life questionnaire. RESULTS: Compared to the TLC diet alone, the TLC diet plus soy nuts was associated with a 45% decrease in hot flashes (7.5 +/- 3.6 vs. 4.1 +/- 2.6 hot flashes day, respectively, p < 0.001) in women with >4.5 hot flashes/day at baseline and 41% in those with

Gender differences in V˙O2 and HR kinetics at the onset of moderate and heavy exercise intensity in adolescents.

The majority of the studies on V˙O2 kinetics in pediatric populations investigated gender differences in prepubertal children during submaximal intensity exercise, but studies are lacking in adolescents. The purpose of this study was to test the hypothesis that gender differences exist in the V˙O2 and heart rate (HR) kinetic responses to moderate (M) and heavy (H) intensity exercise in adolescents. Twenty-one healthy African-American adolescents (9 males, 15.8 ± 1.1 year; 12 females, 15.7 ± 1 year) performed constant work load exercise on a cycle ergometer at M and H. The V˙O2 kinetics of the male group was previously analyzed (Lai et al., Appl. Physiol. Nutr. Metab. 33:107-117, 2008b). For both genders, V˙O2 and HR kinetics were described with a single exponential at M and a double exponential at H. The fundamental time constant (τ1) of V˙O2 was significantly higher in female than male at M (45 ± 7 vs. 36 ± 11 sec, P < 0.01) and H (41 ± 8 vs. 29 ± 9 sec, P < 0.01), respectively. The functional gain (G1) was not statistically different between gender at M and statistically higher in females than males at H: 9.7 ± 1.2 versus 10.9 ± 1.3 mL min-1 W-1, respectively. The amplitude of the slow component was not significantly different between genders. The HR kinetics were significantly (τ1, P < 0.01) slower in females than males at M (61 ± 16 sec vs. 45 ± 20 sec, P < 0.01) and H (42 ± 10 sec vs. 30 ± 8 sec, P = 0.03). The G1 of HR was higher in females than males at M: 0.53 ± 0.11 versus 0.98 ± 0.2 bpm W-1 and H: 0.40 ± 0.11 versus 0.73 ± 0.23 bpm W-1, respectively. Gender differences in the V˙O2 and HR kinetics suggest that oxygen delivery and utilization kinetics of female adolescents differ from those in male adolescents.

Ca2+ as an extracellular signal in bone.

Bone is the major sink and store for calcium and it fulfils essential roles in the maintenance of extracellular free ionised calcium concentration ([Ca2+]e) within its homeostatic range (1.1-1.3 mM). In response to acute hypercalcaemia or hypocalcaemia, Ca2+ is rapidly transported into or out of bone. Bone turnover (and therefore bone Ca2+ turnover) achieves the long-term correction of the [Ca2+]e by the metabolic actions of osteoblasts and osteoclasts, as they respectively incorporate or release Ca2+ from bone. These processes are regulated by the actions of hormones, such as parathyroid hormone (PTH), the release of which is a function of the [Ca2+]e, and is regulated by the action of the Ca2+-sensing receptor (CaR) in the parathyroid gland. Tissue culture studies indicate that bone cells also directly respond to increasing and decreasing [Ca2+]e in their vicinity, independently of the systemic factors. Nevertheless, further studies are necessary to identify how the acute and long-term local changes in [Ca2+]e affect bone cells and the physiological processes they are involved in. Also, the molecular mechanisms which enable the bone cells to sense and respond to [Ca2+]e are not clear. Like the parathyroid cells, bone cells also express the CaR, and accumulating evidence indicates the involvement of this receptor in their responses to the changing extracellular ionic environment.

Osteoblast extracellular Ca2+ -sensing receptor regulates bone development, mineralization, and turnover.

The extracellular Ca(2+) -sensing receptor (CaR), a G protein-coupled receptor responsible for maintenance of calcium homeostasis, is implicated in regulation of skeletal metabolism. To discern the role of the osteoblast CaR in regulation of bone development and remodeling, we generated mice in which the CaR is excised in a broad population of osteoblasts expressing the 3.6-kb a(1) (I) collagen promoter. Conditional knockouts had abnormal skeletal histology at birth and developed progressively reduced mineralization secondary to retarded osteoblast differentiation, evident by significantly reduced numbers of osteoblasts and decreased expression of collagen I, osteocalcin, and sclerostin mRNAs. Elevated expression of ankylosis protein, ectonucleotide pyrophosphatase/phosphodiesterase 1, and osteopontin mRNAs in the conditional knockout indicate altered regulation of genes important in mineralization. Knockout of the osteoblast CaR also resulted in increased expression of the receptor activator of NF-κB ligand (RANKL), the major stimulator of osteoclast differentiation and function, consistent with elevated osteoclast numbers in vivo. Osteoblasts from the conditional knockouts exhibited delayed differentiation, reduced mineralizing capacity, altered expression of regulators of mineralization, and increased ability to promote osteoclastogenesis in coculture experiments. We conclude that CaR signaling in a broad population of osteoblasts is essential for bone development and remodeling and plays an important role in the regulation of differentiation and expression of regulators of bone resorption and mineralization.

Increased oxidative stress in healthy children following an exercise program: a pilot study.

UNLABELLED: Exercise can induce oxidative stress or an imbalance between reactive oxygen species and cellular antioxidant defenses. OBJECTIVE: We investigated the effect of a real-life exercise program on systemic oxidative stress measured by urinary concentrations of 8-isoprostaglandin F2alpha (8-iso-PGF2 alpha), a noninvasive index of lipid peroxidation, in a well-characterized pediatric group. METHODS: Healthy but primarily sedentary, 8- to 10-year-old children (n = 6, mean age 8.8 +/- 0.9 years) of equally distributed healthy weight, overweight, and obese categories, participated in a 5-week exercise program (track and field summer camp, 2 hours/day, 1-2 days/week). RESULTS: By using high-performance liquid chromatography with online electrospray ionization tandem mass spectrometry (LC/ESI/MS/MS), we found a significant (p = .028) increase in group mean urinary 8-iso-PGF2 alpha concentration from 8.163 +/- 6.919 ng/mg creatinine pre-exercise program to 32.320 +/- 16.970 ng/mg creatinine post-exercise program. The increase was also measured at each individual level. We found preliminary evidence that pre- and post-exercise program urinary 8-iso-PGF2 alpha concentrations selectively correlated with children's cardiometabolic characteristics and mood. CONCLUSION: Our results warrant further exploration of the relationships between pre/post-exercise oxidative stress marker 8-iso-PGF2 alpha and cardiometabolic characteristics, exercise habits, eating habits, and mood to determine whether increased post-exercise oxidative stress in healthy children is part of their normal adaptation to exercise or mediator of oxidative injury.

Influence of exercise intensity on pulmonary oxygen uptake kinetics at the onset of exercise and recovery in male adolescents.

The dynamics of the pulmonary oxygen uptake (VO2) responses to square-wave changes in work rate can provide insight into bioenergetic processes sustaining and limiting exercise performance. The dynamic responses at the onset of exercise and during recovery have been investigated systematically and are well characterized at all intensities in adults; however, they have not been investigated completely in adolescents. We investigated whether adolescents display a slow component in their VO2 on- and off-kinetic responses to heavy- and very heavy-intensity exercise, as demonstrated in adults. Healthy African American male adolescents (n=9, 14-17 years old) performed square-wave transitions on a cycle ergometer (from and to a baseline work rate of 20 W) to work rates of moderate (M), heavy (H), and very heavy (VH) intensity. In all subjects, the VO2 on-kinetics were best described with a single exponential at moderate intensity (tau1, on=36+/-11 s) and a double exponential at heavy (tau1, on=29+/-9 s; tau2, on=197+/-92 s) and very heavy (tau1, on=36+/-9 s; tau2, on=302+/-14 s) intensities. In contrast, the VO2 off-kinetics were best described with a single exponential at moderate (tau1, off=48+/-9 s) and heavy (tau1, off=53+/-7 s) intensities and a double exponential at very heavy (tau1, off=51+/-3 s; tau2, off=471+/-54 s) intensity. In summary, adolescents consistently displayed a slow component during heavy exercise (on- but not off-transition) and very heavy exercise (on- and off-transitions). Although the overall response dynamics in adolescents were similar to those previously observed in adults, their specific characterizations were different, particularly the lack of symmetry between the on- and off-responses.

Protein kinase C-mediated phosphorylation of the calcium-sensing receptor is stimulated by receptor activation and attenuated by calyculin-sensitive phosphatase activity.

The agonist sensitivity of the calcium-sensing receptor (CaR) can be altered by protein kinase C (PKC), with CaR residue Thr(888) contributing significantly to this effect. To determine whether CaR(T888) is a substrate for PKC and whether receptor activation modulates such phosphorylation, a phospho-specific antibody against this residue was raised (CaR(pT888)). In HEK-293 cells stably expressing CaR (CaR-HEK), but not in cells expressing the mutant receptor CaR(T888A), phorbol ester (PMA) treatment increased CaR(pT888) immunoreactivity as observed by immunoblotting and immunofluorescence. Raising extracellular Ca(2+) concentration from 0.5 to 2.5 mM increased CaR(T888) phosphorylation, an effect that was potentiated stereoselectively by the calcimimetic NPS R-467. These responses were mimicked by 5 mM extracellular Ca(2+) and abolished by the calcilytic NPS-89636 and also by PKC inhibition or chronic PMA pretreatment. Whereas CaR(T888A) did exhibit increased apparent agonist sensitivity, by converting intracellular Ca(2+) (Ca(2+)(i)) oscillations to sustained plateau responses in some cells, we still observed Ca(2+)(i) oscillations in a significant number of cells. This suggests that CaR(T888) contributes significantly to CaR regulation but is not the exclusive determinant of CaR-induced Ca(2+)(i) oscillations. Finally, dephosphorylation of CaR(T888) was blocked by the protein phosphatase 1/2A inhibitor calyculin, a treatment that also inhibited Ca(2+)(i) oscillations. In addition, calyculin/PMA cotreatment increased CaR(T888) phosphorylation in bovine parathyroid cells. Therefore, CaR(T888) is a substrate for receptor-induced, PKC-mediated feedback phosphorylation and can be dephosphorylated by a calyculin-sensitive phosphatase.

Thiazide diuretics directly induce osteoblast differentiation and mineralized nodule formation by interacting with a sodium chloride co-transporter in bone.

Thiazide diuretics are used worldwide as a first-choice drug for patients with uncomplicated hypertension. In addition to their antihypertensive effect, thiazides increase bone mineral density and reduce the prevalence of fractures. Traditionally, these effects have been attributed to increased renal calcium reabsorption that occurs secondary to the inhibition of the thiazide-sensitive sodium chloride cotransporter (NCC) in the distal tubule. The aim of the current study was to determine whether thiazides exert a direct bone-forming effect independent of their renal action. We found that the osteoblasts of human and rat bone also express NCC, suggesting that these bone-forming cells may be an additional target for thiazides. In vitro, NCC protein was virtually absent in proliferating human and fetal rat osteoblasts, whereas its expression dramatically increased during differentiation. Thiazides did not affect osteoblast proliferation, but directly stimulated the production of the osteoblast differentiation markers runt-related transcription factor 2 (runx2) and osteopontin. Using overexpression/knockdown studies in fetal rat calvarial cells, we show that thiazides increase the formation of mineralized nodules, but loop diuretics do not. Overall, our study demonstrates that thiazides directly stimulate osteoblast differentiation and bone mineral formation independent of their effects in the kidney. Therefore, in addition to their use as antihypertensive drugs, our results suggest that thiazides may find a role in the prevention and treatment of osteoporosis.

WITHDRAWN: Type B gamma-aminobutyric Acid Receptors Modulate the Expression and Function of the Extracellular Ca2+-sensing Receptor in Murine Growth Plate Chondrocytes.

This manuscript was withdrawn at the request of the authors.

Physiological changes in extracellular calcium concentration directly control osteoblast function in the absence of calciotropic hormones.

We investigated the direct effects of changes in free ionized extracellular calcium concentrations ([Ca2+]o) on osteoblast function and the involvement of the calcium-sensing receptor (CaR) in mediating these responses. CaR mRNA and protein were detected in osteoblast models, freshly isolated fetal rat calvarial cells and murine clonal osteoblastic 2T3 cells, and in freshly frozen, undecalcified preparations of human mandible and rat femur. In fetal rat calvarial cells, elevating [Ca2+]o and treatment with gadolinium, a nonpermeant CaR agonist, resulted in phosphorylation of the extracellular signal-regulated kinases 1 and 2, Akt, and glycogensynthase kinase 3beta, consistent with signals of cell survival and proliferation. In agreement, cell number was increased under these conditions. Expression of the osteoblast differentiation markers core binding factor alpha1, osteocalcin, osteopontin, and collagen I mRNAs was increased by high [Ca2+]o, as was mineralized nodule formation. Alkaline phosphatase activity was maximal for [Ca2+]o between 1.2 and 1.8 mM. Inhibition of CaR by NPS 89636 blocked responses to the CaR agonists. In conclusion, we show that small deviations of [Ca2+]o from physiological values have a profound impact on bone cell fate, by means of the CaR and independently of systemic calciotropic peptides.