Yuanzhen Peng

FSH Directly Regulates Bone Mass

Postmenopausal osteoporosis, a global public health problem, has for decades been attributed solely to declining estrogen levels. Although FSH levels rise sharply in parallel, a direct effect of FSH on the skeleton has never been explored. We show that FSH is required for hypogonadal bone loss. Neither FSHbeta nor FSH receptor (FSHR) null mice have bone loss despite severe hypogonadism. Bone mass is increased and osteoclastic resorption is decreased in haploinsufficient FSHbeta+/- mice with normal ovarian function, suggesting that the skeletal action of FSH is estrogen independent. Osteoclasts and their precursors possess G(i2alpha)-coupled FSHRs that activate MEK/Erk, NF-kappaB, and Akt to result in enhanced osteoclast formation and function. We suggest that high circulating FSH causes hypogonadal bone loss.

Diet-induced insulin resistance promotes amyloidosis in a transgenic mouse model of Alzheimer's disease

Recent epidemiological evidence indicates that insulin resistance, a proximal cause of Type II diabetes [a non‐insulin dependent form of diabetes mellitus (NIDDM)], is associated with an increased relative risk for Alzheimers disease (AD). In this study we examined the role of dietary conditions leading to NIDDM‐like insulin resistance on amyloidosis in Tg2576 mice, which model AD‐like neuropathology. We found that diet‐induced insulin resistance promoted amyloidogenic β‐amyloid (Aβ) Aβ1–40 and Aβ1–42 peptide generation in the brain that corresponded with increased γ‐secretase activities and decreased insulin degrading enzyme (IDE) activities. Moreover, increased Aβ production also coincided with increased AD‐type amyloid plaque burden in the brain and impaired performance in a spatial water maze task. Further exploration of the apparent interrelationship of insulin resistance to brain amyloidosis revealed a functional decrease in insulin receptor (IR)‐mediated signal transduction in the brain, as suggested by decreased IR β‐subunit (IRβ) Y1162/1163 autophosphorylation and reduced phosphatidylinositol 3 (PI3)‐kinase/pS473‐AKT/Protein kinase (PK)‐B in these same brain regions. This latter finding is of particular interest given the known inhibitory role of AKT/PKB on glycogen synthase kinase (GSK)‐3α activity, which has previously been shown to promote Aβ peptide generation. Most interestingly, we found that decreased pS21‐GSK‐3α and pS9‐GSK‐ 3β phosphorylation, which is an index of GSK activation, positively correlated with the generation of brain C‐terminal fragment (CTF)‐γ cleavage product of amyloid precursor protein, an index of γ‐secretase activity, in the brain of insulin‐resistant relative to normoglycemic Tg2576 mice. Our study is consistent with the hypothesis that insulin resistance may be an underlying mechanism responsible for the observed increased relative risk for AD neuropathology, and presents the first evidence to suggest that IR signaling can influence Aβ production in the brain.

Cyclooxygenase (COX)-2 and COX-1 potentiate β-amyloid peptide generation through mechanisms that involve γ-secretase activity

In previous studies we found that overexpression of the inducible form of cyclooxygenase, COX-2, in the brain exacerbated β-amyloid (Aβ) neuropathology in a transgenic mouse model of Alzheimers disease. To explore the mechanism through which COX may influence Aβ amyloidosis, we used an adenoviral gene transfer system to study the effects of human (h)COX-1 and hCOX-2 isoform expression on Aβ peptide generation. We found that expression of hCOXs in human amyloid precursor protein (APP)-overexpressing (Chinese hamster ovary (CHO)-APPswe) cells or human neuroglioma (H4-APP751) cells resulting in 10-25 nm prostaglandin (PG)-E2 concentration in the conditioned medium coincided with an ∼1.8-fold elevation of Aβ-(1-40) and Aβ-(1-42) peptide generation and an ∼1.8-fold induction of the C-terminal fragment (CTF)-γ cleavage product of the APP, an index of γ-secretase activity. Treatment of APP-overexpressing cells with the non-selective COX inhibitor ibuprofen (1 μm, 48 h) or with the specific γ-secretase inhibitor L-685,458 significantly attenuated hCOX-1- and hCOX-2-mediated induction of Aβ peptide generation and CTF-γ cleavage product formation. Based on this evidence, we next tested the hypothesis that COX expression might promote Aβ peptide generation via a PG-E2-mediated mechanism. We found that exposure of CHO-APPswe or human embryonic kidney (HEK-APPswe) cells to PG-E2 (11-deoxy-PG-E2) at a concentration (10 nm) within the range of PG-E2 found in hCOX-expressing cells similarly promoted (∼1.8-fold) the generation of the CTF-γ cleavage product of APP and commensurate Aβ-(1-40) and Aβ-(1-42) peptide elevation. The study suggests that expression of COXs may influence Aβ peptide generation through mechanisms that involve PG-E2-mediated potentiation of γ-secretase activity, further supporting a role for COX-2 and COX-1 in Alzheimers disease neuropathology.

TNFα mediates the skeletal effects of thyroid-stimulating hormone

We have shown recently that by acting on the thyroid-stimulating hormone (TSH) receptor (TSHR), TSH negatively regulates osteoclast differentiation. Both heterozygotic and homozygotic TSHR null mice are osteopenic with evidence of enhanced osteoclast differentiation. Here, we report that the accompanying elevation of TNFα, an osteoclastogenic cytokine, causes the increased osteoclast differentiation. This enhancement in TSHR−/− and TSHR+/− mice is abrogated in compound TSHR−/−/TNFα−/− and TSHR+/−/TNFα+/− mice, respectively. In parallel studies, we find that TSH directly inhibits TNFα production, reduces the number of TNFα-producing osteoclast precursors, and attenuates the induction of TNFα expression by IL-1, TNFα, and receptor activator of NF-κB ligand. TSH also suppresses osteoclast formation in murine macrophages and RAW-C3 cells. The suppression is more profound in cells that overexpress the TSHR than those transfected with empty vector. The overexpression of ligand-independent, constitutively active TSHR abrogates osteoclast formation even under basal conditions and in the absence of TSH. Finally, IL-1/TNFα and receptor activator of NF-κB ligand fail to stimulate AP-1 and NF-κB binding to DNA in cells transfected with TSHR or constitutively active TSHR. The results suggest that TNFα is the critical cytokine mediating the downstream antiresorptive effects of TSH on the skeleton.

ACTH protects against glucocorticoid-induced osteonecrosis of bone.

We report that adrenocorticotropic hormone (ACTH) protects against osteonecrosis of the femoral head induced by depot methylprednisolone acetate (depomedrol). This therapeutic response likely arises from enhanced osteoblastic support and the stimulation of VEGF by ACTH; the latter is largely responsible for maintaining the fine vascular network that surrounds highly remodeling bone. We suggest examining the efficacy of ACTH in preventing human osteonecrosis, a devastating complication of glucocorticoid therapy.

The Central Nervous System (CNS)-independent Anti-bone-resorptive Activity of Muscle Contraction and the Underlying Molecular and Cellular Signatures

Background: Mechanisms by which muscle regulates bone are poorly understood. Results: Electrically stimulated muscle contraction reversed elevations in bone resorption and increased Wnt signaling in bone-derived cells after spinal cord transection. Conclusion: Muscle contraction reduced resorption of unloaded bone independently of the CNS, through mechanical effects and, potentially, nonmechanical signals (e.g. myokines). Significance: The study provides new insights regarding muscle-bone interactions. Muscle and bone work as a functional unit. Cellular and molecular mechanisms underlying effects of muscle activity on bone mass are largely unknown. Spinal cord injury (SCI) causes muscle paralysis and extensive sublesional bone loss and disrupts neural connections between the central nervous system (CNS) and bone. Muscle contraction elicited by electrical stimulation (ES) of nerves partially protects against SCI-related bone loss. Thus, application of ES after SCI provides an opportunity to study the effects of muscle activity on bone and roles of the CNS in this interaction, as well as the underlying mechanisms. Using a rat model of SCI, the effects on bone of ES-induced muscle contraction were characterized. The SCI-mediated increase in serum C-terminal telopeptide of type I collagen (CTX) was completely reversed by ES. In ex vivo bone marrow cell cultures, SCI increased the number of osteoclasts and their expression of mRNA for several osteoclast differentiation markers, whereas ES significantly reduced these changes; SCI decreased osteoblast numbers, but increased expression in these cells of receptor activator of NF-κB ligand (RANKL) mRNA, whereas ES increased expression of osteoprotegerin (OPG) and the OPG/RANKL ratio. A microarray analysis revealed that ES partially reversed SCI-induced alterations in expression of genes involved in signaling through Wnt, FSH, parathyroid hormone (PTH), oxytocin, and calcineurin/nuclear factor of activated T-cells (NFAT) pathways. ES mitigated SCI-mediated increases in mRNA levels for the Wnt inhibitors DKK1, sFRP2, and sclerostin in ex vivo cultured osteoblasts. Our results demonstrate an anti-bone-resorptive activity of muscle contraction by ES that develops rapidly and is independent of the CNS. The pathways involved, particularly Wnt signaling, suggest future strategies to minimize bone loss after immobilization.

Regulated production of the pituitary hormone oxytocin from murine and human osteoblasts

Oxytocin (OT) is a primitive neurohypophyseal hormone that plays a primary and indispensible role in mammalian lactation. We have shown recently that OT also regulates bone remodeling, mainly bone formation, with remarkable sensitivity. We now show that OT, apart from its neurohypophyseal origin, is produced in abundance by both human and murine osteoblasts. Production of osteoblast OT is under the control of estrogen, which acts by activating the MAP kinase Erk. This non-genomic mechanism of estrogen action is in stark contrast to its genomic control of OT receptor (OTR) expression. We surmise that there is a local feed-forward loop in bone marrow through which the OT so produced from osteoblasts in response to estrogen acts upon its receptor to exert a potent anabolic action.

Anabolic steroids reduce spinal cord injury-related bone loss in rats associated with increased Wnt signaling

Abstract Background Spinal cord injury (SCI) causes severe bone loss. At present, there is no practical treatment to delay or prevent bone loss in individuals with motor-complete SCI. Hypogonadism is common in men after SCI and may exacerbate bone loss. The anabolic steroid nandrolone reduces bone loss due to microgravity or nerve transection. Objective To determine whether nandrolone reduced bone loss after SCI and, if so, to explore the mechanisms of nandrolone action. Methods Male rats with complete transection of the spinal cord were administered nandrolone combined with a physiological replacement dose of testosterone, or vehicle, beginning on day 29 after SCI and continued for 28 days. Results SCI reduced distal femoral and proximal tibial bone mineral density (BMD) by 25 and 16%, respectively, at 56 days. This bone loss was attenuated by nandrolone. In ex vivo osteoclasts cultures, SCI increased mRNA levels for tartrate-resistant acid phosphatase (TRAP) and calcitonin receptor; nandrolone-normalized expression levels of these transcripts. In ex vivo osteoblast cultures, SCI increased receptor activator of NF-kB ligand (RANKL) mRNA levels but did not alter osteoprotegerin (OPG) mRNA expression; nandrolone-increased expression of OPG and OPG/RANKL ratio. SCI reduced mRNA levels of Wnt signaling-related genes Wnt3a, low-density lipoprotein receptor-related protein 5 (LRP5), Fzd5, Tcf7, and ectodermal-neural cortex 1 (ENC1) in osteoblasts, whereas nandrolone increased expression of each of these genes. Conclusions The results demonstrate that nandrolone reduces bone loss after SCI. A potential mechanism is suggested by our findings wherein nandrolone modulates genes for differentiation and activity of osteoclasts and osteoblasts, at least in part, through the activation of Wnt signaling.

Oxytocin deficiency impairs maternal skeletal remodeling.

We have reported that the posterior pituitary hormone, oxytocin (OT), known for its effects in inducing parturition, lactation and social bonding, is also a skeletal hormone. Here, we demonstrate that OT plays a key role in enabling maternal skeletal mobilization during pregnancy by enhancing the formation of bone resorbing osteoclasts. Osteoclast formation ex vivo is thus diminished in pregnant mothers with genetic OT-deficiency. OT(-/-) pups at day E20 also show a defect in trabecular bone. microCT measurements reveal normal bone volume, but increased trabecular numbers, suggesting that trabeculae in OT(-/-) pups are hypomineralized. We suggest that OT facilitates intergenerational transfer of calcium ions from a pregnant mother to the pups.

Nandrolone slows hindlimb bone loss in a rat model of bone loss due to denervation

Nandrolone is an anabolic steroid that has been demonstrated to reduce the loss of bone and muscle from hindlimb unweighting and to slow muscle atrophy after nerve transection. To determine whether nandrolone has the ability to protect bone against loss due to disuse after denervation, male rats underwent sciatic nerve transaction, followed 28 days later by treatment with nandrolone or vehicle for 28 days. Bone mineral density (BMD) was determined 28 days later or 56 days after nerve transection. Denervation led to reductions in BMD of 7% and 12% for femur and tibia, respectively. Nandrolone preserved 80% and 60% of BMD in femur and tibia, respectively, demonstrating that nandrolone administration significantly reduced loss of BMD from denervation. This study offers a potential novel pharmacological strategy for use of nandrolone to reduce bone loss in severe disuse‐ and denervation‐related bone loss, such as that which occurs after spinal cord injury.