Search

The dynamic behavior of the thin plates resting on a fractionally damped viscoelastic foundation subjected to a moving point load is investigated. This work explores the application of fractional c...

The dynamic behaviour of an Euler–Bernoulli beam resting on the fractionally damped viscoelastic foundation subjected to a moving point load is investigated. The fractional-order derivative-based Kelvin–Voigt model describes the rheological properties of the viscoelastic foundation. The Riemann–Liouville fractional derivative model is applied for a fractional derivative order. The modal superposition method and Triangular strip matrix approach are applied to solve the fractional differential equation of motion. The dependence of the modal convergence on the system parameters is studied. The influences of (a) the fractional order of derivative, (b) the speed of the moving point load and (c) the foundation parameters on the dynamic response of the system are studied and conclusions are drawn. The damping of the beam-foundation system increases with increasing the order of derivative, leading to a decrease in the dynamic amplification factor. The results are compared with those using the classical integer-order derivative-based foundation model. The classical foundation model over-predicts the damping and under-predicts the dynamic deflections and stresses. The results of the classical (integer-order) foundation model are verified with literature.

The dynamic response spectra of fractionally damped viscoelastic beams subjected to concentrated moving load are presented. This work quantitatively emphasizes the need for fractional-order damping...

The free vibration analysis of thin orthotropic plates with elastically constrained edges is presented using the energy-based Rayleigh–Ritz (RR) method. Various edge conditions are modeled with rotational and translational linear springs. The complete set of admissible functions, which is a combination of (1) trigonometric functions, (2) the unit function, (3) the linear function, and (4) the lowest order polynomial, has been used in the RR method. It has been demonstrated that the use of a combination of the lowest order polynomial and the cosine series results in a rapid convergence of the solution, without any ill-conditioning of the admissible functions upon expansion. In particular, this work proposes a simple guideline for determining a set of trial functions that can be universally utilized for the vibration analysis of plates with non-classical boundary conditions. The convergence and exactness of this approach have been demonstrated through several examples. The results indicate that the elastically restrained stiffness and the plate aspect ratio impact the frequency parameters considerably.

The coupled horizontal-torsional-warping vibration of a thin-walled open-section 7800 TEU container ship bare-hull, modelled as a non-uniform girder, is analysed by the efficient energy-based Rayleigh-Ritz method, in order to generate the dry asymmetric vibration frequencies. Since the centre of gravity is within the hull and the shear centre is below the keel, the horizontal and torsional modes of vibration are highly coupled. An open section is also prone to warping. In a novel attempt, the bare-hull geometry is generated mathematically, using section-wise closed-form semi super-ellipses (Lame’s curves). The main dimensions, weight distributions, and fineness ratios are preserved, and closed-form expressions of sectional properties become available in the process. The hull has arbitrarily (non-mathematically) varying mass, bending stiffness, warping stiffness, and shear stiffness distributions along the length. The non-uniform beam modeshape in horizontal/torsional vibration is assumed to be a weighted sum of the uniform beam horizontal/torsional modeshapes. Several benchmark cases of simpler geometry have been analysed first, for both torsion-warping vibration, and coupled horizontal-torsional-warping vibration. Pontoon approximation of the containership has been analysed and validated. Subsequently, the coupled dry vibration frequencies are obtained for the open deck non-uniform girder, and compared with published results.

This work underlines the importance of the application of fractional-order derivative damping model in the modelling of the viscoelastic foundation, by demonstrating the effect of various orders of the fractional derivative on the dynamic response of plates resting on the viscoelastic foundation, subjected to concentrated step load. The foundation of the plate is modelled as a fractionally-damped Kelvin–Voigt model. Modal superposition method and Triangular strip matrix approach are used to solve the partial fractional differential equations of motion. The influence of (a) fractional-order derivative, (b) foundation stiffness, and (c) foundation damping viscosity parameter on the dynamic response of the plate are investigated. Theoretical results show that with the increase in the order of derivative, the damping of the system increases, which leads to decreased dynamic response. The results obtained from the fractional-order damping model and integer-order damping model are compared. The results are verified with literature and numerical results (ANSYS).

Viscoelastic materials (VEM) are often used in the ship floors to dampen the noise and vibration induced by machinery (engines, pumps, alternators) and live loads (human traffic and liquid movement in pipelines). Therefore, accurate modeling of such materials is important for the iterative design-analysis process. VEM exhibits both time and frequency-dependent properties. Traditionally, integer-order viscoelastic mechanical models are used to describe the rheological properties of the viscoelastic materials. However, such models find difficulties in predicting the rheological characteristics since their kernel functions are a combination of exponential functions. For instance, the integer-order Maxwell model is good at describing the stress relaxation behavior while is poor in capturing creep, and vice-versa for the integer-order Kelvin-Voigt model. However, the fractional-order mechanical model can overcome the above problem with a fewer number of model parameters. Therefore, in this paper, the fractional derivative-based Kelvin-Voigt mechanical model is employed to describe the time-dependent vibratory behavior of the VEM. To validate the effectiveness of the above model, a thin elastic plate bonded with a thin layer of VEM, subject to a concentrated impact load, is studied. Galerkin’s method and Triangular strip matrix approach are used to solve the partial fractional differential equations of motion. The semi-analytical approach of modal superposition is used to generate the response, whose first dynamic overshoot of displacement is crucial to contain the dynamic stress within the working stress level. This bypasses the computationally expensive Finite Element Method. Additionally, ANSYS is limited to the integer-order damping model only. This analysis gives insights into the efficacy of the VEM chosen for passive vibration control of structural components of ship hulls. A case study for free vibration is done with a commercially available VEM, which is used as surface flooring to cover steel plates of ship hulls, thereby acting as vibration dampers. Free vibration results show that the damping coefficient of the plate foundation system increases with increasing the order of the derivative. In addition, the amplitude of the transient response decreases with the order of the derivative. Thus, the classical integer-order mechanical model overestimates the damping of the viscoelastic materials, which leads to underestimating the displacement and associated stresses. The results are verified with literature and ANSYS.

A semi-analytical approach to free dry and wet vibration of a trapezoidal, 2-way tapered, pivoted hollow spade rudder is presented. The rudder is modeled as a hollow Kirchhoff’s plate, with a NACA0018 profile chord section. The rudder pivot is modeled as a combination of a translational spring and a rotational spring. The span-wise and chord-wise non-uniform beam vibration is first analyzed by the Rayleigh-Ritz method, to establish the non-uniform beam modeshapes, which act as admissible functions to the Galerkin’s method for plate vibration. Eigenvalue analysis generates the plate natural frequencies and the plate modeshapes. Alternately, uniform beam modeshapes themselves are used as admissible functions into the Galerkin’s method. Frequencies are analyzed for various pivot positions, taper ratios, and NACA sections. For the wet vibration, constant strength source distribution technique is used to generate the added mass of a 2D aerofoil. Also, 3D panel method is used to generate the modal added masses, and hence the wet natural frequencies. The added mass coefficient is generated for various aerofoil fineness ratios, pivot fixities, taper ratios, aspect ratios.

The dynamic response of fractionally damped viscoelastic plates subjected to a moving point load is investigated. In order to capture the viscoelastic dynamic behavior more accurately, the material is modeled using the fractionally damped Kelvin–Voigt model (rather than the integer-type viscoelastic model). The Riemann–Liouville fractional derivative of order 0 < α ≤ 1 is applied. Galerkin's method and Newton–Raphson technique are used to evaluate the natural frequencies and corresponding damping coefficients. The structure is subject to a moving point load, traveling at different speeds. The modal summation technique is applied to generate the dynamic response of the plate. The influence of the order of the fractional derivative on the free and transient vibrations is studied for different velocities of the moving load. The results are compared with those using the classical integer-type Kelvin–Voigt viscoelastic model. The results show that an increase in the order of the fractional derivative causes a significant decrease in the maximum dynamic amplification factor, especially in the “dynamic zone” of the normalized sweep time. The dynamic behavior of the plate is verified with ansys.

Parathyroid hormone-related peptide (PTHrP) is well-known to play a role in bone formation, and abaloparatide, an analog of PTHrP(1-34), is approved for the treatment of osteoporosis in post-menopausal women. PTHrP has also been reported to have cardiovascular effects, with recent data demonstrating that exogenously administered PTHrP can limit the death of isolated cardiomyocytes subjected to oxidative stress via upregulation of classic ‘survival kinase’ signaling. Our aim in the current study was to extend this concept and, employing both in vitro and in vivo models, establish whether PTHrP(1-36) and abaloparatide are cardioprotective in the setting of lethal myocardial ischemia-reperfusion injury. We report that preischemic administration of PTHrP(1-36) and abaloparatide attenuated cell death in HL-1 cardiomyocytes subjected to simulated ischemia-reperfusion, an effect that was accompanied by the augmented expression of phospho-ERK and improved preservation of phospho-Akt, and blocked by co-administration of the MEK-ERK inhibitor PD98059. Moreover, using the translationally relevant swine model of acute coronary artery occlusion-reperfusion, we make the novel observation that myocardial infarct size was significantly reduced in pigs pretreated with PTHrP(1-36) when compared with placebo-controls (13.1 ± 3.3% versus 42.0 ± 6.6% of the area of at-risk myocardium, respectively; p < 0.01). Taken together, these data provide the first evidence in support of the concept that pretreatment with PTHrP(1-36) and abaloparatide renders cardiomyocytes resistant to lethal myocardial ischemia-reperfusion injury.

The aim is to analyze the wave-induced vertical vibration of a non-prismatic mathematical hull in a stochastic sea, by normal mode analysis. The hull has been generated mathematically, to represent two distinct Indian merchant vessels: DS (Tanker) and SCIM (Containership). The body-plan, deck waterline, bow and stern profiles, have been modeled as semi-superellipses. These render non-uniform distributions of mass and stiffness over the ship-length. The energy-based Rayleigh–Ritz method has been used to analyze the idealized hull girder natural frequencies and modeshapes. The non-uniform beam modeshape is a weighted series sum of prismatic beam-free vibration modeshapes. The 2D added mass of superelliptic sections is formulated, solving the radiation boundary value problem by the constant strength source distribution method. The hull girder is subject to the Pierson–Moskowitz sea spectrum in fully loaded condition. The diffraction force is formulated through the Khaskind’s relations. The flexural response of the girder is evaluated by the modal superposition method. The response spectra have been generated for various sea states and ship speeds. The magnitudes of the maximum flexural/shear stress for each vessel are generated. The probability of shear/tensile failure is also estimated, giving insights into the hull structural design.

A compliant tower is modeled as a partially dry, partially tapered, damped Timoshenko beam with the superstructure modeled as an eccentric tip mass, and a non-classical damped boundary at the base. The foundation is modeled as a combination of a linear spring and a torsional spring, along with parallel linear and torsional dampers (Kelvin-Voigt model). The superstructure adds to the kinetic energy of the system without affecting the potential energy, thereby reducing the natural frequencies. The weight of the superstructure acts as an axial compressive load on the beam, reducing its natural frequencies further. The empty space factor due to the truss-type structure of the tower is included. The effect of shear deformation and rotary inertia are included in the vibration analysis; with the non-uniform beam mode-shapes being a weighted sum of the uniform beam mode-shapes satisfying the end condition. The weights are evaluated by the Rayleigh-Ritz (RR) method, and verified using finite element method (FEM). The weight of the superstructure acts as an axial compressive load on the beam. Kelvin-Voigt model of structural damping is included. A part of the structure being underwater, the virtual added inertia is included to calculate the wet natural frequencies. A parametric study is done for various magnitudes of tip mass and various levels of submergence. The computational efficiency and accuracy of the Rayleigh-Ritz method, as compared to the FEA, has been demonstrated. The advantage of using closed-form trial functions is clearly seen in the efficacy of calculating the various energy components in the RR method.

A closed-form approach is proposed to scrutinize and study the prominence of rigid-body modes (RBM) of a translationally edge-restrained thin plate. Accurate vibrational analysis of an elastically edge-supported plate is not possible without the inclusion of degenerate rigid-body beam-wise admissible functions. The trivial solution(s) of the beam frequency equation produce(s) a non-trivial modeshape, which satisfies the boundary conditions, has zero curvature, and is orthogonal to other modeshapes. The frequency parameters, waveform coefficients, orthogonality, and potential energies of the RBM have been studied for a wide range of edge restraints, asymptotically leading to the classical edge beam vibration characteristics. The second trivial frequency parameter is seen to transform from a RBM to a flexural mode at a certain range of edge spring constant. The first several (20) non-trivial modeshapes of a fully free (FFFF) square plate have been semi-analytically generated and are seen to match well with published experimental work. The nature of the frequencies has been scrutinized using Eigenvectors, with respect to the participation of the RBM functions, which are seen to dominate the plate modeshape even in higher-order frequencies. However, they are mutually exclusive: the translational RBM influences the odd flexural modes, while the rotational RBM influences the even flexural modes. The confusions in literature, especially about the identical frequencies, have been clarified.

Dynamic analysis of thin rectangular elastically supported plates to transient loads is presented. A floating airport is modeled as a horizontal Kirchhoff’s plate, which is elastically supported at the ends, and is subjected to the impact of aircrafts landing and deceleration over its length. This sets the free–free–free–free plate into high-frequency vibration, causing flexural stress waves to travel over the plate. First, the beam natural frequencies and modeshapes in either direction are generated with these complexities. The eigenvalue analysis of the governing differential equation is done, using the weighted summation of the product of the beam modes. The radiation pressure on the bottom side of the plate is included to reduce the frequencies by the added-mass effect. The plate is then subjected to decelerating shock loads. The vibratory response is analyzed by the computationally efficient normal mode analysis. The amplification factor versus the taxiing time of the moving load is generated. This gives insights into the maximum stress encountered under the transient load, as function of taxiing time and support.

The free vibration of a low-aspect cantilever wing is studied by semianalytical, numerical, and experimental means. The wing is modeled as a two-way tapered, hollow Kirchhoff's plate, with the chord-wise section as symmetrical NACA0018 aerofoil. The chord length and the thickness taper from root to tip, over the span. The semianalytical approach is based on Galerkin's method, which includes the modal superposition of two orthogonal beam modeshapes (free-free beam in chord-wise direction and cantilever beam in span-wise direction). The free vibration is also studied numerically using ANSYS. The results have been compared with an experimental study, performing the dry impact hammer test. A model scale wing has been constructed from a 3-mm-thick metal sheet, with a length-scale ratio of 1:10. Comparative studies have been done among the three methods. The feasibility of modeling a low-aspect-ratio wing as a plate has been investigated. 1. Introduction Lifting surfaces commonly occur in engineering structures: fans, steam and gas turbine blades, impeller blades, wind turbine blades, helicopter fans, airplane wings, marine propeller blades, marine rudders and skegs, and other control surfaces like hydrofoils and wings in high-speed marine crafts. The lifting surface is typically a cantilever, with one edge welded to the main structure (root), and the other end free (tip). The span-wise geometry is tapered, for greater strength at the root and lighter weight at the tip. The chord-wise cross section is an aerofoil, which acts as a lifting surface to the incoming flow at varying angles of attack. The aspect ratio of the lifting surface, i.e., the ratio of the span length to the mean chord length, may vary from 0.4 to 0.6 in a turbine blade to 12 to 15 in an airplane wing/wind turbine. This work focuses on a low-aspect-ratio lifting surface, whose usual span-to-chord ratio is 1.5–2.

An as-yet limited body of evidence suggests that calcium-regulating endocrine hormones—in particular, parathyroid hormone–related peptide (PTHrP)—may have unappreciated cardioprotective effects. The current review focuses on the concept that PTHrP may, via modulation of classic cardioprotective signaling pathways, provide a novel strategy to attenuate myocardial ischemia–reperfusion injury.

A semi-analytical approach to understand the manifestation of plate modeshapes associated with twin frequencies has been presented. Square Mindlin’s plate, clamped on all sides, has been considered here. It highlights the importance of efficacy of the beam-wise trial functions in an energy-based plate vibration analysis method, in terms of (a) accuracy, (b) orthogonality, (c) sense (plus/minus) and (d) interference. The inconsistency in the modeshapes of repeated frequencies, seen extensively in literature, has been attempted to be removed, through superior closed-form orthogonal set of Timoshenko admissible functions into the Rayleigh-Ritz method. The constructive/destructive interferences of the admissible functions, which are the products of the beam-wise modeshapes, give the final nodal patterns and the prominence of the anti-nodes. Also, the pairs of ‘very close’ but distinct frequencies, which were often considered as ‘numerical errors’, have been counter-intuitively justified through their Eigenvectors, which are either symmetric or skew-symmetric in the matrix form. Nodal patterns for CCCC plate modeshapes are accurately investigated; i.e. chess-board and diagonal nodal patterns.

A semianalytical study of a uniform homogenous partially submerged square cantilever plate vibration is presented. The structure is assumed to be a Kirchhoff's plate, clamped on one edge and free on the other edges. The lengthwise section of the plate is a cantilever clamped-free (CF) beam, while the widthwise section is a free-free (FF) beam. The plate modeshape is a weighted superposition of the product of the beam modeshapes, with unknown weights. The CF beam has only flexural modes. The FF beam has two rigid-body modes, i.e., translational and rotational modes. Rayleigh–Ritz method (RRM) is used to set up the free vibration eigenvalue problem. The eigenvector gives the unknown weights. The modeshapes generated are further used in the boundary element method (BEM) to calculate the fluid inertia, which participates in the vibration and leads to a consistent drop in frequencies. The dependence of this reduction on the submergence level is studied for the first six frequencies of the plate. The frequencies are also experimentally generated by the impact hammer test, both in the dry state, and under three distinct levels of submergence: 25%, 50%, and 75% from the free edge opposite to the clamped edge. The frequencies and modeshapes are also verified through numerical analysis using the commercial code ansys 16.0. Conclusions are drawn regarding the influence of fluid inertia distribution on the final plate modeshape, leading to insights into sound structural designs.

A theoretical hydroelastic analysis of an axially loaded uniform Timoshenko beam, with intermediate end fixities, undergoing hydrodynamic impact-induced bottom slamming, is presented. The underwater part of a marine craft is modeled as a lightly damped flexible beam, which rises out of the water in rough seas, and slams against it at a very large vertical velocity. This causes highly intense, localized hydrodynamic impact pressure sweeping across the beam at high velocities, setting it into high-frequency vibrations, predisposing it to plastic deformation and/or fatigue failure. The natural frequencies of the structure depend on the slenderness ratio, axial load, and end fixities. The natural frequencies and modeshapes are generated through the Eigen analysis. The changing wetted surface is the prime complexity of the problem. The relative velocity between fluid and structure is considered to establish the hydrodynamic pressure (radiation and impact). Normal mode summation method is used to analyze the transient structural vibration, for various impact speeds, deadrise angles, end fixities, and axial loads on the beam. The primary aim is to establish the zone of prominence of hydroelasticity, study the maximum dynamic stress magnitudes under various loading conditions and structural parameters; and draw conclusions leading to insights into sound structural designs.

Previous attempts to solve for vibration of non-uniform hull girder have used Finite Element Analysis (FEA). This work utilizes the Rayleigh–Ritz (R–R) method to analyze the vertical vibration of a non-prismatic mathematical hull, with arbitrary longitudinal distribution of sectional properties. It is shown that (a) the R-R method provides reasonably accurate results for the actual natural frequency; and (b) the R–R method offers significant computational advantages over FEA. This computational supremacy can be exploited in the initial design stages when several designs are to be iteratively tested for its structural characteristics. The natural frequencies and modeshapes are obtained by the Rayleigh–Ritz method. The non-uniform beam modeshape is a weighted series sum of the (closed-form) uniform beam modeshapes. Only a few uniform beam modes are necessary to generate the non-uniform beam modeshape and a convergent hull girder frequency. Thus, the method is very effective in the low-frequency domain like ship-structure. The proposed method is compared with FEA for two illustrative structures : (a) a containership and (b) a tanker; and also with several hulls from established literature. Modal convergence studies have also been included. The distortions of the non-uniform modeshapes have been studied in the light of the loading conditions of the hull.

The free vibration of a continuous, elastic model of a mono-piled ocean tower is studied. The tower, partially submerged in water, is undergoing free transverse vibration in a plane. It is modeled as a non-uniform Timoshenko beam which has an eccentric tip mass on one end and is supported by a mono-piled foundation on the other. Effects of shear deformation and rotary inertia are included in the beam. The problem of interaction of soil with the pile is solved by adopting the Winkler Foundation model. The viscoelastic character of the soil is included using the Kelvin–Voigt model. Equation of motion of the tower is derived using Hamilton׳s variational principle and approximate analytical solution is found using the Rayleigh Ritz Method (RRM). The solution is compared with that obtained from a conventional Finite Element Method (FEM) which shows a good agreement. The trial function of RRM is assumed as uniform beam mode-shapes satisfying the boundary and continuity conditions of the ocean tower. At the end, an extensive parametric study is carried out which provides an insight into the dependence of natural frequency on different configurations of the tower.

In this review, we have highlighted work that has clearly demonstrated that mitogen-activated protein kinase (MAPK) phosphatase-1 (MKP-1), a negative regulator of MAPKs, is an important signaling mediator in bone, muscle, and fat tissue homeostasis and differentiation. Further, we examined recent studies with particular focus on MKP-1 overexpression or deletion and its impact on tissues connected to bone. We also summarized regulation of MKP-1 by known skeletal regulators like parathyroid hormone (PTH)/PTH-related peptide (PTHrP) and bone morphogenic proteins. MKP-1's integration into the pathophysiological state of osteoporosis, osteoarthritis, rheumatoid arthritis, obesity, and muscular dystrophy are examined to emphasize possible involvement of MKP-1 both at the molecular level and in disease complications such as sarcopenia- or diabetes-related osteoporosis. We predict that understanding the mechanism of MKP-1-mediated signaling in bone-muscle-fat crosstalk will be a key in coordinating their activities and developing therapeutics to improve clinical outcomes for diseases associated with advanced age.

This paper studies a continuous, elastic model of an ocean tower, partially submerged in water, undergoing free transverse vibration in a plane. The tower is modeled as a non-uniform Timoshenko beam which is supported by an eccentric tip mass on one end and a non-classical damped foundation on the other. The foundation is modeled as a combination of translational and rotational springs and dampers. The effect of shear deformation and rotary inertia is included in the analysis. The free vibration equation is derived using Hamilton׳s variational principle based on two approaches, Rayleigh Ritz Method (RRM) and Finite Element Method (FEM), which show a good agreement in results. The computational efficiency of RRM over FEM is shown using a convergence study. Finally, a parametric study is done to demonstrate the dependence of natural frequency on different configurations of the tower.

The free wet vibration characteristics of an idealized low-aspect-ratio cantilever wing are studied semi-analytically, numerically, and experimentally. The wing is modeled as a tapered, hollow Kirchhoff’s plate, with the chord-wise section as a symmetrical NACA0018 aerofoil. The chord length tapers from root to tip, over the span. The main aim is to set up a suitable radiation boundary problem for the vibrating cantilever wing, in order to semi-analytically generate the wet frequencies. The efficacy of the semi-analytical wet vibration approach is studied by comparing it with the other two approaches. The difficulties encountered are due to the hollow two-way tapered shape of the wing and the free edge boundary conditions on its three sides. The semi-analytical approach is based on Galerkin’s method, which includes the modal superposition of two orthogonal beam modeshapes (Free-Free beam in chord-wise and cantilever in span-wise directions). The plate modeshapes thus generated are further used in the 3D source distribution technique to calculate the fluid inertia, leading to a consistent drop in the natural frequencies. The cantilever wing has been fabricated and tested in-house. The underwater impact hammer test generates the wet natural frequencies. The free vibration frequencies are also verified numerically using ANSYS, and compared with experimental studies.

Bone mass is dependent on osteoblast proliferation, differentiation and life-span of osteoblasts. Parathyroid hormone (PTH) controls osteoblast cell cycle regulatory proteins and suppresses mature osteoblasts apoptosis. Intermittent administration of PTH increases bone mass but the mechanism of action are complex and incompletely understood. Cell Cycle and Apoptosis Regulatory Protein (CARP)-1 (aka CCAR1) is a novel transducer of signaling by diverse agents including cell growth and differentiation factors. To gain further insight into the molecular mechanism, we investigated involvement of CARP-1 in PTH signaling in osteoblasts. Immunostaining studies revealed presence of CARP-1 in osteoblasts and osteocytes, while a minimal to absent levels were noted in the chondrocytes of femora from 10 to 12-week old mice. Treatment of 7-day differentiated MC3T3-E1 clone-4 (MC-4) mouse osteoblastic cells and primary calvarial osteoblasts with PTH for 30min to 5h followed by Western blot analysis showed 2- to 3-fold down-regulation of CARP-1 protein expression in a dose- and time-dependent manner compared to the respective vehicle treated control cells. H-89, a Protein Kinase A (PKA) inhibitor, suppressed PTH action on CARP-1 protein expression indicating PKA-dependent mechanism. PMA, a Protein Kinase C (PKC) agonist, mimicked PTH action, and the PKC inhibitor, GF109203X, partially blocked PTH-dependent downregulation of CARP-1, implying involvement of PKC. U0126, a Mitogen-Activated Protein Kinase (MAPK) Kinase (MEK) inhibitor, failed to interfere with CARP-1 suppression by PTH. In contrast, SB203580, p38 inhibitor, attenuated PTH down-regulation of CARP-1 suggesting that PTH utilized an Extracellular Signal Regulated Kinase (ERK)-independent but p38 dependent pathway to regulate CARP-1 protein expression in osteoblasts. Immunofluorescence staining of differentiated osteoblasts further revealed nuclear to cytoplasmic translocation of CARP-1 protein following PTH treatment. Collectively, our studies identified CARP-1 for the first time in osteoblasts and suggest its potential role in PTH signaling and bone anabolic action.

We examined core body temperature (CBT) of urocortin 1 (UCN1) and corticotropin releasing factor (CRF) knockout (KO) mice exposed to 4°C for 2 h. UCN1KO mice showed higher average CBT during cold exposure as compared to WT. The CBT of male and female WT mice dropped significantly to 34.1 ± 2.4 and 34.9 ± 3.1 C at 4°C, respectively. In contrast, the CBT of male and female UCN1KO mice dropped only slightly after 2 h at 4°C to 36.8 ± 0.7 and 38.1 ± 0.5 C, respectively. WT female and male UCN1KO mice showed significant acclimatization to cold; however, female UCN1KO mice did not show such a significant acclimatization. CRFKO mice showed a dramatic decline in CBT from 38.2 ± 0.4 at 22°C to 26.1 ± 9.8 at 4°C for 2 h. The CRF/UCN1 double KO (dKO) mice dropped their CBT to 32.5 ± 4.0 after 2 h exposure to 4°C. Dexamethasone treatment prevented the decline in CBT of the CRFKO and the dKO mice. Taken together, the data suggest a novel role for UCN1 in thermoregulation. The role of CRF is likely secondary to adrenal glucocorticoids, which have an important regulatory role on carbohydrate, fat, and protein metabolism.

Limited information is available on the role of MAPK phosphatase1 (MKP1) signaling in osteoblasts. We have recently reported distinct roles for MKP1 during osteoblast proliferation, differentiation and skeletal responsiveness to parathyroid hormone (PTH). Since MKP1 regulates the phosphorylation status of MAPKs we investigated the involvement of P-ERK and P-p38 MAPKs in MKP1 knock out (KO) early and mature osteoblasts with respect to mineralization and PTH response. Calvarial osteoblasts from 9–14 week old wild type (WT) and MKP1 KO male and female mice were examined. Western blot analysis revealed down-regulation and sustained expressions of P-ERK and P-p38 with PTH treatment in differentiated osteoblasts derived from KO males and females respectively. Exposure of early osteoblasts to p38 inhibitor, SB203580 (S), markedly inhibited mineralization in WT and KO osteoblasts from both genders as determined by von Kossa assay. In osteoblasts from males, ERK inhibitor U0126 (U), not p38 inhibitor (S), prevented the inhibitory effects of PTH on mineralization in early or mature osteoblasts. In osteoblasts from KO females, PTH sustained mineralization in early osteoblasts and decreased mineralization in mature cells. This effect of PTH was attenuated by S in early osteoblasts, and by U in mature KO cells. Changes in matrix gla protein (MGP) expression with PTH in KO osteoblasts did not correlate with mineralization, indicative of MKP1 dependent additional mechanisms essential for PTH action on osteoblast mineralization. We conclude that PTH regulation of osteoblast mineralization in female mice is maturation stage specific and involves MKP1 modulation of P-ERK and P-p38 MAPKs.

Activation of G protein-coupled receptors by agonists leads to receptor phosphorylation, internalization of ligand receptor complexes, and desensitization of hormonal response. The role of parathyroid hormone (PTH) receptor 1, PTHR1, is well characterized and known to regulate cellular responsiveness in vitro. However, the role of PTHR1 phosphorylation in bone formation is yet to be investigated. We have previously demonstrated that impaired internalization and sustained cAMP stimulation of phosphorylation-deficient (PD) PTHR1 leads to exaggerated cAMP response to subcutaneous PTH infusion in a PD knockin mouse model. To understand the physiological role of receptor internalization on PTH bone anabolic action, we examined bone parameters of wild-type (WT) and PD knockin female and male mice following PTH treatment. We found a decrease in total and diaphyseal bone mineral density in female but not in male PD mice compared with WT controls at 3–6 mo of age. This effect was attenuated at older age groups. PTH administration displayed increased bone volume and trabecular thickness in the vertebrae and distal femora of both WT and PD animals. These results suggest that PTHR1 phosphorylation does not play a major role in the anabolic action of PTH.

Phosphorylation, internalization, and desensitization of G protein-coupled receptors, such as the parathyroid hormone (PTH) and PTH-related peptide (PTHrP) receptor (PTH1R), are well characterized and known to regulate the cellular responsiveness in vitro. However, the role of PTH1R receptor phosphorylation in bone formation and osteoblast functions has not yet been elucidated. In previous studies, we demonstrated impaired internalization and sustained cAMP stimulation of a phosphorylation-deficient (pd) PTH1R in vitro, and exaggerated cAMP and calcemic responses to s.c. PTH infusion in pdPTH1R knock-in mouse model. In this study, we examined the impact of impaired PTH1R phosphorylation on the skeletal phenotype of mice maintained on normal, low, and high calcium diets. The low calcium diet moderately reduced (P1/S phase cyclin, in vitro. In contrast to WT osteoblasts, down-regulation of cyclin D1 was sustained for longer periods of time in osteoblasts isolated from the pd mice. Our results suggest that adaptive responses of intracellular signaling pathways in the pd mice may be important for maintaining bone homeostasis.

Although there is a growing interest in the molecular cross-talk between the endocrine and cardiovascular systems, the cardiac effects of calcium-regulating hormones (i.e., parathyroid hormone-related peptide (PTHrP)) have not been explored. In this study, we examined the effect of PTHrP on the viability of isolated adult mouse cardiomyocytes subjected to oxidative stress. Myocytes from 19 to 22 week old male 129J/C57BL6 mice were exposed to oxidative insult in the form of H2O2 which led to more than 70% loss of cell viability. Herein we demonstrate, for the first time, that pretreatment with 100 nM PTHrP prior to 100 μM H2O2 incubation prevents H2O2 -induced cell death by more than 50%. Immunoblot analysis revealed H2O2 induction of MKP-1 protein expression while PTHrP decreased MKP-1 expression. Moreover, myocytes derived from MKP1 KO mice were resistant to oxidative injury. No added benefit of PTHrP treatment was noted in MKP-1 null cardiomyocytes. Using specific pharmacological inhibitors we demonstrated that P-p38, P-ERK and P-AKT mediated PTHrP's cardioprotective action. These data provide novel evidence that: i) down-regulation of MKP1 affords profound protection against oxidative stress; and ii) PTHrP is cardioprotective, possibly via down-regulation of MKP-1 and activation of MAPK and PI3K/AKT signaling.

A theoretical analysis of free dry and wet vibration of a trapezoidal, 2-way tapered, marine spade rudder, is presented. The rudder is considered as a hollow Kirchhoff’s plate, with the chord section as a NACA profile. The chord length and the thickness taper from the top to the bottom, over the vertical span. The rudder is pivoted at the top, with the pivot behind the leading edge. The pivot is modeled as a combination of a translational and a rotational spring, in order to include the rigid body modes of the rudder vibration. The span-wise and chord-wise non-uniform beam vibration is first analyzed by the Rayleigh-Ritz method, in order to establish the non-uniform beam mode shapes. The span-wise beam is a linearly tapered vertical cantilever, with non-classical edge at the top and free at the bottom. The chord-wise section is a 2-span beam with the ends free, and four continuity conditions at the pivot. The non-uniform mode shapes, in either direction, are a weighted summation of the uniform beam mode shapes, which also satisfy the boundary/continuity conditions. They now act as admissible spatial functions to the plate vibration, which is analyzed by the Galerkin’s method. Eigenvalue analysis generates the plate natural frequencies. A weighted superposition, of the product of the beam mode shapes, in either direction, generates the plate mode shapes. Alternately, uniform beam mode shapes are used as admissible functions into the Galerkin’s method for the plate natural frequencies and mode shapes. The natural frequencies are generated for various positions of the rudder stock along the chord length. The pivot conditions (in both translational and rotational rigid body degree of freedom) influence the prominence of the rigid body mode shapes. The natural frequencies are analyzed for various pivot fixities, taper ratios, and aspect ratios of the plate. This is followed by the wet vibration analysis of the rudder. First, 2D strip theory is used to generate the added mass of each chord section. Constant strength source distribution technique is used to generate the added mass in sway and yaw of a 2D aerofoil. Each flexural and torsional mode is associated with its own added mass. Various empirical corrections are done to account for the 3D flow. Finally, 3D panel method is used to generate the modal added masses, and hence the wet natural frequencies. The added mass coefficient is generated for various aerofoil fineness ratios, pivot fixities, taper ratios, and aspect ratios of the plate.

Rap1, a growth regulatory protein that is strongly expressed in human squamous cell carcinoma (SCC), is inactivated by rap1GAP. Recent evidence in normal rat cells suggests that rap1GAP regulates proliferation. The objective of the current study was to in-vestigate whether rap1GAP functions as a tumor suppressor in SCC. Using a pull-down assay, active GTP-bound rap1 was up-regulated in SCC compared to normal or immortalized keratinocytes. Because both rap1A and rap1B isoforms of rap1 are expressed in SCC, the rap1GAP inactivation of both rap1 isoforms was verified using cells transfected with EGFP-rap1A or EGFP-rap1B or co-transfected with FLAG-tagged rap1GAP. The results demonstrate that expression of rap1GAP in oropharyngeal SCC down-regulated active rap1, ERK activation, and proliferation. Incubation of stably transfected SCC cells with nocodazole, an inhibitor of mitosis, caused a slower accumulation of rap1GAP-transfected cells in the G2 phase, in comparison to the vector control, indicating that rap1GAP-transfected cells have slower progression through the cell cycle. This was supported by down-regulation of cyclin D1, cdk4, and cdk6 in rap1GAP-transfected SCC cells. Furthermore, SCC cells transfected with rap1GAP produced significantly smaller tumors in nude mice as compared to controls (P < 0.01). These novel findings suggest that rap1GAP acts as a tumor suppressor protein in SCC.

PTH is an anabolic agent used to treat osteoporosis, but its mechanisms of action are unclear. This study elucidated target cells and mechanisms for anabolic actions of PTH in mice duringbonegrowth.Micewithc-fosablationareosteopetrotic and lack an anabolic response to PTH. In this study, there were no alterations in PTH-regulated osteoblast differentiation or proliferation in vitro in cells from c-fos / mice compared with /; hence, the impact of osteoclastic cells was further investigated. A novel transplant model was used to rescue the osteopetrotic defect of c-fos ablation. Vertebral bodies (vossicles) from c-fos/and/mice were implanted into athymic hosts, and the c-fos / osteoclast defect was rescued. PTH treatment to vossicle-bearing mice increased 5-bromo-2-deoxyuridine (BrdU) positivity in the bone marrow and increased bone area regardless of the vossicle genotype. To inhibit recruitment of osteoclast precursors to wildtypevossicles,stromalderivedfactor-1signalingwasblocked, which blunted the PTH anabolic response. Treating mice with osteoprotegerin to inhibit osteoclast differentiation also blocked the anabolic action of PTH. In contrast, using c-src mutant mice with a late osteoclast differentiation defect did not hinder the anabolic action, suggesting key target cells reside in the intermediately differentiated osteoclast population in the bone marrow. These results indicate that c-fos in osteoblasts is not critical for PTH action but that cells of the osteoclast lineage are intermediate targets for the anabolic action of PTH. (Endocrinology 146: 4584–4596, 2005)

Objective This study was undertaken to address the involvement of CDK activating kinase (CAK), p53, and MDM2 proteins in the mitotic arrest associated with the acquisition of a polyploid DNA content during megakaryocyte differentiation of human erythroleukemia (HEL) cells. Methods To evaluate this mechanism we investigated HEL cells as a model system in which there is a marked increase in DNA content during megakaryocyte differentiation induced by phorbol-diesters. Specific cell-cycle phases were separated by centrifugal elutriation and SDS PAGE and Western analysis were performed to determine the relative abundance of these proteins. Kinase assays were carried out following immunoprecipitation of cellular lysates with the antibodies to the proteins. Results Polyploid HEL cells show an increase in the abundance of the CAK complex proteins, CDK7 and cyclin H, and a sixfold increase in CAK-specific activity. Increased CAK activity in polyploid HEL cells follows both the downregulation of p53 protein and its decreased association with CAK complex. Consistent with the reduction of p53, polyploid HEL cells undergo a dramatic increase in MDM2 protein abundance that in turn facilitates increased interaction of this protein with p53. Conclusion These observations demonstrate that deregulated expression of MDM2 and p53 during megakaryocyte differentiation allow a relaxation of the control over genomic stability, allowing further replicative rounds of DNA synthesis.

There is growing interest in the molecular cross-talk between the endocrine and cardiovascular systems, including the effects of hormones on cardiomyocte viability in models of ischemic and oxidative stress. However, current attention has focused on reproductive and metabolic hormones; the cardiac effects of calcium-regulating hormones (i.e., parathyroid hormone-related peptide (PTHrP)) have not been explored. To address this, we interrogated the effect of PTHrP on the viability of isolated adult mouse cardiomyocytes subjected to H 2 O 2 -induced oxidative stress. In Aim 1 , myocytes from wild type (WT) C57 mice were incubated for 16 hrs with 100 nM PTHrP or vehicle and exposed to 100 μM H 2 O 2 for 15 min. Myocyte viability was significantly improved in PTHrP-treated cells vs vehicle control (70±12% vs 20±15%; mean ± SD, pAim 2 ), we: (i) incubated WT cardiomyocytes with PTHrP (100 nM) and probed for expression of MKP1; and (ii) repeated the viability assay as described above using myocytes from adult MKP1 knockout (KO) mice. Incubation of WT myocytes with PTHrP evoked a 55% decrease in MKP1 expression. Moreover, myocytes from MKP1 KO mice were resistant to oxidative injury: viability was maintained at 78±12% in vehicle controls (mean ± SD, pvs the value of 20±15% in vehicle-treated WT cells), with no added benefit of PTHrP treatment (Figure). Finally ( Aim 3 ), in both models (PTHrP treatment; MKP1 KO), the enhanced cell viability was accompanied by ~2-fold increases in Akt phosphorylation. These data provide novel evidence that: i) down-regulation of MKP1 affords profound protection against oxidative stress; and ii) PTHrP is cardioprotective, possibly via down-regulation of MKP1 and activation of Akt signaling.

The free vibration of a rudder is studied here by theoretical, numerical, and experimental means. The rudder is modeled as a Kirchhoff’s plate, with chord-wise sections as symmetrical aerofoils of NACA0018 section. The free vibration is also studied numerically. The results have been compared with experimental studies. A model scale rudder has been constructed from a 3 mm thick metal sheet, with a length-scale ratio of 1:10. Impact hammer test has been down to excite all the natural frequencies, which have been picked up by two accelerometers into an FFT analyzer. Comparative studies have been done among the three methods.

A compliant tower (CT) is modeled as a partially dry, partially tapered, damped Timoshenko beam with the superstructure modeled as an eccentric tip mass, and a non-classical damped boundary at the base. The foundation is modeled as a combination of a linear spring and a torsional spring, along with linear and torsional dampers. The mean empty space factor due to the truss type structure of the tower is included. The effect of shear deformation and rotary inertia are included in the vibration analysis; with the non-uniform beam mode-shapes being a weighted sum of the uniform beam mode-shapes. The weights are evaluated by the Rayleigh-Ritz method, using the first ten modes and verified using Finite Element Method (FEM). The superstructure adds to the kinetic energy without affecting the stiffness of the beam, thereby reducing the natural frequencies. The weight of the superstructure acts as an axial compressive load on the beam, reducing its frequencies further. Kelvin-Voigt model of structural damping is included. A part of the structure being underwater, the virtual added inertia is included to calculate the wet natural frequencies. The CT is first subjected to steady current loads of a given velocity profile. The static deflection and overturning moment is estimated for current loads. The CT is then studied for wave excitation at various seas states. Morrison’s equation and Pierson-Moskowitz Spectrum are used to derive the forces for different sea states. The forced vibration analysis of the structure is done via Rayleigh-Ritz method and verified using FEM. The maximum horizontal deflection and shear stress of the base of the superstructure, and the normal/shear stresses at the foundation are analyzed. Finally, the CT is subjected to earthquake excitation, modeled as an arbitrary horizontal impact excitation at the base. The above forced vibration analysis is repeated.Copyright © 2014 by ASME

The role of MAPKs in osteogenesis and in Parathyroid hormone (PTH) signaling has been shown in vitro and in vivo previously. However, the role of MAPK phosphatase-1 (MKP-1) in bone formation remains unclear. We observed age dependent sexual dimorphism in bone phenotype of MKP-1 knock out (KO) mice showing osteopenia in females and osteopetrosis in males. We investigate the involvement of MKP-1 on b-catenin, a key mediator of the canonical Wnt signaling pathway and an important regulator of bone forming cells, the osteoblasts. Signal transduction through b-catenin requires inhibition of GSK-3b which is also inhibited via phosphorylation by AKT upon insulin-like growth factor-1 (IGF-1) signaling. Real-time PCR analysis of primary calvarial osteoblasts revealed significant down-regulation of IGF-1 mRNA expression in osteoblasts isolated from MKP-1 KO female mice in contrast to KO osteoblasts from males. Inhibition of P-ERK and P-p38 with pharmacological inhibitors showed disparate regulation of IGF-1 express...

Little is known concerning the interaction of thrombopoietin (TPO) with other megakaryocyte-active cytokines in directing the early events of megakaryocyte development. Culture of CD34+ cells in interleukins (IL) -1, -6, -11, plus stem cell factor (SCF; S) results in a 10- to 12-fold expansion in total cell numbers, whereas total CD41+ megakaryocytes are expanded ∼120-fold over input levels. Addition of TPO to IL-1, -6, -11, S generates a biphasic proliferation of CD41+ cells, accelerates their rate of production, and results in an ex vivo expansion of more than 200-fold. The addition of Flt-3 ligand (FL) increases CD41+ cell expansion to ∼380-fold over input levels. In the absence of TPO, ∼95% of the expanded cells show the phenotype of promegakaryoblasts; TPO and/or FL addition increases CD41 antigen density and ploidy in a subpopulation of promegakaryoblasts. A moderate (approximately sevenfold) expansion of megakaryocyte progenitor cells (colony-forming unit-megakaryocyte) occurs in the presence of IL-1, -6, -11, S, and the addition of TPO to this cocktail yields an ∼17-fold expansion. We conclude that early proliferative events in megakaryocyte development in vitro are regulated by multiple cytokines, and that TPO markedly affects these early developmental steps. However, by itself, TPO is neither necessary nor sufficient to generate a full proliferative/maturational in vitro response within the megakaryocyte compartment. TPO clearly affects terminal differentiation and the development of (some) high-ploidy human megakaryocytes. However, its limited in vitro actions on human cell polyploidization suggest that additional megakaryocyte-active cytokines or other signals are essential for the maximal development of human megakaryocytes.

Metabolic bone disease occurs when there is a net loss in bone density. Osteoporosis, the most common metabolic bone disease, is a devastating problem and an increasingly major public health issue. A substantial body of evidence in the elderly population indicates that a relationship exists between the components of body weight and various measures of bone/mass, density, and function. Both muscle and fat contribute to the body’s total weight and the intimate associations of muscle, fat, and bone are known. But the close functional interactions between muscle and bone or fat and bone are largely unidentified and have drawn much attention in recent years. Each of these tissues not only responds to afferent signals from traditional hormone systems and the central nervous systems but also secretes factors with important endocrine functions. Studies suggest that during growth, development, and aging, the relationship of muscle and fat with the skeleton possibly governs bone homeostasis and turnover. A better understanding of the endocrine function and the cellular and molecular mechanisms and pathways linking muscle or adipose tissues with bone anabolism and catabolism is a new avenue for novel pathways for anabolic drug discovery. These in turn will likely lead to more rational therapy toward increasingly prevalent disorders like osteoporosis. In this review, some of the recent works on the interaction of bone with muscle and fat are highlighted, and in so doing the role of parathyroid hormone (PTH), and PTH-related peptide (PTHrP) is surveyed.

A dynamic analysis of axially loaded plates with intermediate fixities is presented. The underwater part of a craft is modeled as a lightly damped flexible plate, which rises out of the water and slams against it at a large vertical velocity. This causes highly localized hydrodynamic impact pressure moving at high velocities across the plate, setting it into high-frequency vibrations. The natural frequencies of the plate depend on the slenderness ratio, aspect ratio, axial load, and end fixities. The end fixities depend on the quality of welding, and type of bracketing. Normal mode summation method is used to analyze the vibratory response, which is generated for various impact speeds, deadrise angles, end fixities, axial loads, aspect ratios, and slenderness ratios. A parametric study is done to predict the location, instant, and magnitude of the maximum dynamic stresses on the structure. The hydrodynamic pressure is assumed to act on the equilibrium position of the plate, i.e. one-way coupling of the fluid and the structure is studied. The primary aim is to study the time-varying dynamic stress configurations and draw conclusions leading to sound structural designs.Copyright © 2013 by ASME

Dynamic analysis of thin rectangular elastically supported stiffened plates with axial loads is presented. A floating airport is modeled as a horizontal Kirchhoff’s plate, which is elastically supported at the ends; and is subjected to the impact of aircrafts landing and deceleration over its length. This sets the free-free-free-free plate into high-frequency vibration, causing flexural stress waves to travel over the plate. First, the beam natural frequencies and modeshapes in either direction are generated with these complexities. The Eigen value analysis of the governing differential equation is done, using the weighted summation of the product of the beam modes. The accuracy of the frequencies is compared with those from FEA studies. The radiation pressure on the bottom side of the plate is included to reduce the frequencies by the added-mass effect. The plate is then subjected to decelerating shock loads. The vibratory response is analyzed by the computationally efficient normal mode analysis. The amplification factor vs. the taxiing time of the moving load is generated.Copyright © 2013 by ASME

Parathyroid hormone (PTH) signaling via PTH 1 receptor (PTH1R) involves mitogen-activated protein kinase (MAPK) pathways. MAPK phosphatase 1 (MKP1) dephosphorylates and inactivates MAPKs in osteoblasts, the bone-forming cells. We previously showed that PTH1R activation in differentiated osteoblasts upregulates MKP1 and downregulates pERK1/2–MAPK and cyclin D1. In this study, we evaluated the skeletal phenotype ofMkp1knockout (KO) mice and the effects of PTHin vivoandin vitro. Microcomputed tomography analysis of proximal tibiae and distal femora from 12-week-oldMkp1KO female mice revealed osteopenic phenotype with significant reduction (8–46%) in bone parameters compared with wild-type (WT) controls. Histomorphometric analysis showed decreased trabecular bone area in KO females. Levels of serum osteocalcin (OCN) were lower and serum tartrate-resistant acid phosphatase 5b (TRAP5b) was higher in KO animals. Treatment of neonatal mice with hPTH (1–34) for 3 weeks showed attenuated anabolic responses in the distal femora of KO mice compared with WT mice. Primary osteoblasts derived from KO mice displayed delayed differentiation determined by alkaline phosphatase activity, and reduced expressions ofOcnandRunx2genes associated with osteoblast maturation and function. Cells from KO females exhibited attenuated PTH response in mineralized nodule formationin vitro. Remarkably, this observation was correlated with decreased PTH response of matrix Gla protein expression. Expressions of pERK1/2 and cyclin D1 were inhibited dramatically by PTH in differentiated osteoblasts from WT mice but much less in osteoblasts fromMkp1KO mice. In conclusion, MKP1 is important for bone homeostasis, osteoblast differentiation and skeletal responsiveness to PTH.

Osteoporosis and age-related bone loss is associated with changes in bone remodeling characterized by decreased bone formation relative to bone resorption, resulting in bone fragility and increased risk of fractures. Stimulating the function of bone-forming osteoblasts, is the preferred pharmacological intervention for osteoporosis. Recombinant parathyroid hormone (PTH), PTH(1-34), is an anabolic agent with proven benefits to bone strength and has been characterized as a potential therapy for skeletal repair. In spite of PTH’s clinical use, safety is a major consideration for long-term treatment. Studies have demonstrated that intermittent PTH treatment enhances and accelerates the skeletal repair process via a number of mechanisms. Recent research into the molecular mechanism of PTH action on bone tissue has led to the development of PTH analogs to control osteoporotic fractures. This review summarizes a number of advances made in the field of PTH and bone fracture to combat these injuries in humans and in animal models. The ultimate goal of providing an alternative to PTH, currently the sole anabolic therapy in clinical use, to promote bone formation and improve bone strength in the aging population is yet to be achieved.

Parathyroid hormone (PTH) and PTH-related protein (PTHrP) activate one single receptor (PTH1R) which mediates catabolic and anabolic actions in the bone. Activation of PTH1R modulates multiple intracellular signaling responses. We previously reported that PTH and PTHrP down-regulate pERK1/2 and cyclin D1 in differentiated osteoblasts. In this study we investigate the role of MAPK phosphatase-1 (MKP-1) in PTHrP regulation of ERK1/2 activity in relation to osteoblast proliferation, differentiation and bone formation. Here we show that PTHrP increases MKP-1 expression in differentiated osteoblastic MC3T3-E1 cells, primary cultures of differentiated bone marrow stromal cells (BMSCs) and calvarial osteoblasts. PTHrP had no effect on MKP-1 expression in proliferating osteoblastic cells. Overexpression of MKP-1 in MC-4 cells inhibited osteoblastic cell proliferation. Cell extracts from differentiated MC-4 cells treated with PTHrP inactivate/dephosphorylate pERK1/2 in vitro; immunodepletion of MKP-1 blocked the ability of the extract to dephosphorylate pERK1/2; these data indicate that MKP-1 is involved in PTHrP-induced pERK1/2 dephosphorylation in the differentiated osteoblastic cells. PTHrP regulation of MKP-1 expression is partially dependent on PKA and PKC pathways. Treatment of nude mice, bearing ectopic ossicles, with intermittent PTH for 3weeks, up-regulated MKP-1 and osteocalcin, a bone formation marker, with an increase in bone formation. These data indicate that PTH and PTHrP increase MKP-1 expression in differentiated osteoblasts; and that MKP-1 induces growth arrest of osteoblasts, via inactivating pERK1/2 and down-regulating cyclin D1; and identify MKP-1 as a possible mediator of the anabolic actions of PTH1R in mature osteoblasts.

Parathyroid hormone-related protein (PTHrP) is an integral mediator of physiologic and pathologic processes and has demonstrated actions in the periodontium. PTHrP functions via AP-1, and specifically through JunB. This study identified JunB-dependent downstream mediators of PTHrP using OCCM cementoblastic transfectants with JunB over- or reduced expression. Over-expressing cells showed an increase in proliferation, while the opposite was seen in siRNA transfected cells. Microarray analysis of over-expressing cells revealed more than 1000 regulated genes. Three genes were investigated in more detail. The PTH/PTHrP receptor (PTHR1) and ephrin B1 (EfnB1) were down-regulated, and vascular cell adhesion molecule-1 (VCAM-1) was up-regulated with JunB over-expression. JunB siRNA transfectants had increased PTHR1, but reduced ephrin B1 and unaltered VCAM-1 in vitro. To validate these targets, parental OCCM cells and primary osteoblasts were treated with PTHrP, resulting in reduced PTHR1 and ephrin B1, and increased VCAM-1. Cell transfectants were implanted subcutaneously in vivo, and microarray analysis and RT-PCR performed. Over-expression of JunB down-regulated PTHR1 and ephrin B1, and increased VCAM-1. JunB siRNA transfectant implants had increased PTHR1 and ephrin B1, but no altered VCAM-1. These data highlight new gene targets for PTHrP and indicate JunB is a critical mediator of PTHrP actions.