LEPTIN AND BONE
Central control of bone metabolism by leptin
Shu Takeda
Department of Orthopedics and 21COE, Tokyo Medical and Dental University, Tokyo, Japan
Abstract: Leptin, following the binding to ObRb in the hypothalamus, affects bone formation and resorption through multiple pathways. Through the sympathetic nervous system, leptin inhibits bone formation via CREB, AP-1 and the molecular clock, while stimulates bone resorption via ATF4. Moreover, leptin also inhibits bone resorption via CART through an unidentified mechanism.
Key words: bone formation, bone resorption, hypothalamus, sympathetic nervous system, p blocker, cocaine- and amphetamine-regulated transcript
1. I N T R O D U C T I O N
With the increase in longevity, osteoporosis is now emerging as a major concern for public health, since it is the most common degenerative bone disease in the developed world\ Osteoporosis is caused by an imbalance between bone formation and bone resorption^. Clinically, the most frequent cause for osteoporosis is menopause, on the other hand, obese individuals are relatively protected from the development of osteoporosis^. These observations suggest that there may be a link between fat mass and bone mass. Furthermore, as osteoblasts and adipocytes originate from the same mesenchymal progenitor cells^, many investigators searched for molecules linking fat and bone biology. Recently, our laboratory discovered the existence of hypothalamic control of bone remodeling'*, a discovery that was confirmed subsequently by others. Leptin regulates bone formation and resorption via the hypothalamus and sympathetic nervous system^.
Considering that most of leptin's physiological actions are mediated by the hypothalamus*, the notion that central control of bone metabolism by leptin seems to be quite reasonable. However, leptin has been shown to directly
affect cells localized in bone in some reports. Before discussing the central nature of leptin in bone metabolism, I would like to summarize leptin's local effect on bone.
2. LOCAL ACTION OF LEPTIN ON BONE CELL
Leptin is not only produced by visceral or subcutaneous adipocytes, but from bone marrow adipocytes^. Several reports indicate that leptin affects bone cells locally. Leptin functional receptors ObRb are expressed in osteoblasts*'', osteoclasts and chondrocytes'"'", cells indispensable for bone modeling and remodeling. Interestingly, osteoblasts produce and secrete leptin*, suggesting the autocrine nature of leptin in bone remodeling.
Moreover, exogenous leptin increased osteoblastic differentiation or mineralization*'''*^ in vitro, thus demonstrating an anabolic action on bone metabolism. On the contrary, others reported that leptin induced apoptosis of bone marrow stromal cells '^. Those direct effects of leptin were not confined to osteoblasts. Namely, leptin inhibited osteoclast differentiation*'' or stimulated chondrocyte proliferation". In vivo, intraperitoneal treatment of leptin stimulated bone growth of ob/ob mice*^ and reduced ovariectomy- induced bone loss**. These results suggest that leptin affects bone cells in local microenvironment. However, leptin overexpression in bone using bone-specific al(I) collagen promoter does not affect bone metabolism in vivo^. The reason for this discrepancy can be partly explained by the fact that most in vitro studies were performed using supraphysiological amounts of leptin. Taken together, it is indicated that the primary action of leptin in vivo is not mediated by direct action on bone cells.
3. EFFECT OF LEPTIN ON BONE FORMATION THROUGH CENTRAL NERVOUS SYSTEM
Since most of the physiological condition are under control of the central nervous system, especially hypothalamus, one may hypothesize that bone remodeling does not escape this rule. As a molecule explaining the bone sparing effect of obesity, we focused on leptin, which many known actions result following its binding to specific receptors on hypothalamic neurons and whose deficiency causes morbid obesity. Mouse models of leptin deficiency (ob/ob) and leptin receptor-deficiency (db/db) demonstrated a marked high bone mass phenotype'*. This was quite unexpected, since there is no other mouse model in which hypogonadism and high bone mass
phenotypes co-exist. Subsequent studies of the ob/ob mice showed an increased bone formation and resorption, suggesting that high bone mass is due to enhanced bone formation. Intracerebroventricular infusion of leptin results in bone loss in ob/ob mice and wild-type mice without raising serum concentration of leptin, thus established antiosteogenic function of leptin via the hypothalamus^CFigure 1).
^ s>
Leptin Hypothalamus
Sympathetic nervous system AdrP2,^0^
CART
osteoblast
CREB ATF4 / \ \ AP-1 I — Clock RANKL
\ / c-myc
i
cyclinDI
^ \ "
Osteoblast proliferation, Bone formation
osteoclast
Bone resorption
Figure I. Model of central control of bone metabolism by leptin.
Leptin inhibits osteoblast proliferation via sympathetic nervous system. CREB, AP-1 and Clock are involved in this regulation. Leptin also affects bone resorption through osteoclast differentiation factor RANKL.
The melanocortin pathway has a crucial role in leptin's regulation of food intake*. However, genetic (Agouti yellow mice and melanocortin 4 receptor- deficient mice) or pharmacological (MC4R agonist) manipulation of melanocortin pathway did not affect the antiosteogenic i.e. inhibition of bone formation action of leptin^, suggesting that pathways regulating antiosteogenic and anorexigenic function of leptin are different. In the diverse phenotypic abnormality observed in ob/ob mice, low tonus of sympathetic abnormality caught our attention. Namely, we tested if leptin's antiosteogenic function is mediated via sympathetic nervous system. Indeed, dopamine-P-hydroxylase (DBH)-deficient mice, which are unable to synthesize catecholamines and mice treated with non-selective |3 blocker (propranolol) displayed the same high bone mass phenotype than ob/ob
mice, Moreover, the fact that icv infusion of leptin decreased body weight in DBH-deficient mice or propranolol-treated mice, but not their bone mass, demonstrated the essential role of sympathetic nervous system in antiosteogenic action of leptin^ not in its control of body weight (Figure 1).
This observation raises the prospect that blockade of the sympathetic nervous system could be potentially a novel therapy for osteoporosis.
Subsequent analysis of mice lacking the adrenergic P2receptor, the only adrenergic receptor expressed in osteoblasts, revealed high bone mass phenotype accompanied by an increase in bone formation'^, as expected.
More recently, circadian genes were discovered to act as downstream signaling molecules of sympathetic signaling in osteoblasts'*. Indeed, mice lacking molecular clock components displayed high bone mass and showed a paradoxical increase in bone formation following leptin icv infusion. In osteoblasts, leptin and sympathetic nervous system induced clock genes expression, which antagonizes AP-1 family-mediated osteoblast proliferation. Thus, the molecular mechanism of leptin's antiosteogenic action partly, if not all, is now being clarified'*( ure 1).
4. CENTRAL EFFECT ON OSTEOCLASTS BY LEPTIN AND SNS
Surprisingly, adrb2-deficient mice developed another bone phenotype, which is a decrease in bone resorption". Leptin icv infusion caused an increase in bone resorption in wild-type mice, but not in adrb2-deficient mice, demonstrating that leptin increases bone resorption via the sympathetic nervous system. Isoproterenol, a (3 receptor agonist, favored osteoclastic bone resorption by inducing the expression of the osteoclast differentiation factor RANKL in osteoblasts while no effect of sympathetic signaling on osteoclasts could be found. This effect of sympathetic signaling on RANKL expression required the osteoblast specific transcription factor ATF4 phosphorylation by PKA. Importantly, adrb2-deficent mice are protected from gonadectomy-induced bone loss, indicating that sympathetic nervous system is indispensable for the development of postmenopausal osteoporosis. These results demonstrated that leptin regulates both axis of bone remodeling centrally'^ and gave further importance to this regulation since very few molecules regulate both aspects of bone remodeling (Fig. 1).
5. CENTRAL EFFECT ON OSTEOCLASTS BY CART
The increase in bone resorption observed in ob/ob mice was interpreted to be secondary to hypogonadism initially. However, the dissociation of bone resorption status in ovariectomized-b2adr-deficient mice and ob/ob mice argued against that. Namely, though both mutant mouse strains display a hypogonadism and a decrease in sympathetic tone, the former model displayed normal bone resorption, while the latter showed increased bone resorption. These results suggested that the action of leptin on bone resorption is not solely dependent on sympathetic nervous system and led to the identification of CART as another molecule implicated in leptin's action on bone resorption*^. CART is a gene whose expression is regulated by leptin and is nearly absent in ob/ob mice that was thought to be anorectic".
However, CART-deficient mice have a normal food intake, are lean on normal diet^". In contrast, CART-deficient mice are osteopenic due to an isolated increase in osteoclastic resorption'^. Leptin icv caused a further reduction of bone loss compared to wild type animal, suggesting that CART mediates leptin's action modulating bone resorption. In addition, high bone mass phenotype observed in MC4R-deficient mice and human patients can be explained by their increase in CART expression. Taken together these data establish that , leptin regulates bone resorption centrally, through two different pathways, sympathetic nervous system-ATF4-RANKL and
C A R T ' ^
6. L E P T I N , P B L O C K E R S A N D O S T E O P O R O S I S IN C L I N I C A L M E D I C I N E
Clinically, the association of serum leptin concentration and bone mass or bone metabolic markers are under intensive investigation. However, results of various studies are conflicting. Some reports showing no correlation between serum leptin level and bone metabolic markers in postmenopausal women^''^^, while positive correlation was observed between serum leptin level and bone mass in women^^'^^. On the contrary, studies including adult men show that serum leptin level was inversely correlated with bone formation markers and bone mineral density^* or bone mineral density^^. These results might suggest a gender specific effect of leptin in bone metabolism. However, more importantly, the association of leptin and other parameters are lost or diminished after adjusting body weight in most studies. It is well accepted that obesity leads to leptin- resistance, as hyperinsulinemia does in type2 diabetes patients. This is best
demonstrated by the fact that serum leptin concentration and body weight is positively correlated^*. Given the fact that a clinical marker reflecting leptin's action in vivo is not available, studies exploring leptin and bone mineral density in human beings need to be carefully interpreted.
A few studies were conducted to address if leptin treatment affects bone mass in human patients. Farroqi et. al. reported the decrease of bone mineral density after leptin replacement therapy to a single leptin-deficient patient^'.
It has also been shown that leptin supplementation to 2 lipodystrophic patients did not affect bone mass^". Clearly, larger clinical trials are necessary to address the role of leptin on bone metabolism in human beings.
Although the most interesting aspect of this regulation is the involvement of a sympathetic tone which is more amenable to therapeutic manipulations.
Since blockade of sympathetic regulation of bone remodeling increases bone mass in mouse, clinical researchers examined if a P blocker, a most commonly prescribed drug, may be beneficial for bone metabolism in human. Some prospective studies reported the lack of association between P blocker usage and bone density'*'''^, while two large cohort studies found almost 30% reduction of bone fracture by propranolol^''^'*. Recently, a randomized trial assessing the effect of propranolol on bone metabolic markers reported no beneficial effect of a P blocker. However, considering that this study was conducted in a relatively short period with a small number of patients a larger randomized study is warranted.
7. C O N C L U S I O N
Leptin has become, in relatively few years, a major regulator of bone metabolism, whose mode of action has been largely elucidated'^. Recently, various neuropeptides and their receptors, such as N P Y ' * , M C H ' ^ and cannabinoid '*, were identified as regulator of bone metabolism, thus our understanding of central control of bone metabolism is expanding. However, several questions remain to be elucidated. One is obviously the molecular mechanism how CART affects bone resorption. In addition, there may be some other molecules mediating leptin-dependent sympathetic regulation of bone formation. From the clinical point of view, modulating leptin and its downstream pathway, especially bone-specific P blockers, is a potential therapeutic target for the treatment of osteoporosis.
ACKNOWLEDGEMENTS
I thank Gerard Karsenty for critical reading of the manuscript. This worlc was supported in part by grants from Ministry of Education, Culture, Sports, Science, Japan, grants from Center of Excellence Program for Frontier Research on Molecular Destruction and Reconstruction of Tooth and Bone, Japan Society for Promotion of Science, Tokyo, grants from Ono Medical Research Foundation, and grants from Kanae Foundation for Life and Socio-Medical Science.
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