Mechanobiology

The osteocyte is a long-lived, highly dendritic cell forming a vast, interconnected network within the mineralized bone matrix. It has only been over the last couple of decades that cell isolation and imaging techniques have advanced enough to allow deeper study of this most interesting cell. The osteocyte is acknowledged as integral to sensation of changing mechanical forces at the whole bone level (mechanosensation) and has also been identified as the major source of key proteins that modulate osteoblast and osteoclast response to loading (cells that build and resord bone, respectively). However, the mechanisms by which mechanosensation translates to changes in modulating proteins (mechanotransduciton) as well as how these proteins are transported from the osteocyte, embedded within the bone matrix, to the effector cells at the surface and in the marrow space of the bone is poorly understood.

Past work of the BBL has identified unique properties in osteocytes, including robust intracellular Ca²⁺ oscillations that change in frequency and magnitude with increasing mechanical force, and a dense actin cytoskeleton that contracts with the onset of these calcium spikes. Our current work focuses on identifying the precise mechanism of these Ca²⁺-dependent contractions as well as uncovering downstream consequences of this phenomenon. The BBL has developed novel in vitro, ex vivo, and in vivo systems that focus on different aspects of potential mechanisms to help us answer these questions.

Please click on links below for more in depth descriptions of current projects underway in the lab.

Single Cell Studies and Cytoskeleton Mechanics in Quasi-3D

Downstream consequences of Ca2+-mediated contractions in Osteocytes

We have previously demonstrated Ca²⁺ oscillations in osteocytes in response to mechanical loading both in vitro and ex vivo. In addition, mechanical loading is known to regulate osteocyte protein expression. Yet, no studies to date have connected Ca²⁺-mediated mechanosensitivity to protein expression changes in osteocytes, and few studies have linked Ca²⁺ signaling to long-term adaptive responses.

Visualization and characterization of Ca2+-dependent actomyosin contractions in ex vivo osteocytes

Osteocytes exhibit many unique characteristics which have been thought to play a role in their mechanosensing capabilities, including longevity, a unique stellate morphology, and their existence as a vast, interconnected 3D network within the bone matrix. As such, it is important to maintain these features when investigating osteocyte responses to mechanical load. Thus, we have developed an ex vivo model of osteocyte mechanotransduction in which we visualize and capture real-time, short term responses of osteocytes to mechanical loading in explanted mice tibiae.

Influence of Sclerostin antibody on mechanotransduction in ex vivo osteocytes

The BBL’s ex vivo system can be used to evaluate mechanosensitivity of bone, which may be impaired in bone diseases, such as osteoporosis, or affected by therapeutic interventions. One of the most promising treatments for bone loss is the monoclonal antibody against sclerostin (Scl-Ab), which targets a protein produced primarily by osteocytes. Administration of antibodies to sclerostin has been shown to increase bone mineral density by increasing bone formation and decreasing bone resorption in both animal studies and human clinical trials.

Monitoring of formation/resorption events over time using sequential in vivo microcomputed tomography

Osteocytes are known to be the orchestrators of mechanosensation, as evidenced by our in vitro and ex vivo work, but the ultimate downstream effects of this sensation is best assessed using in vivo models. Daily cyclic mechanical loading can be applied to one tibia of a mouse via customized attachments to a load frame typically over 2 to 4 weeks. Often the contralateral limb serves as a non-loaded control. Each week both tibiae are scanned via micro-computed tomography, providing standard morphological measurements of trabecular and cortical bone.

Differentiating modeling and remodeling events using patterns of formation/resorption to differentiate the stimuli of these unique responses

Modeling and remodeling are two unique and essential processes in bone metabolism. Remodeling occurs throughout life, and is the coupled repair process of bone resorption followed by bone formation in the same location. Modeling is an uncoupled formative or resorptive response that is typical of growth or osteoporosis, respectively. The underlying mechanisms of bone mechanobiology are inherently linked to understanding these distinct processes. Furthermore the efficacy of treatments to address bone deficiencies may benefit from understanding which of these processes they affect.

In vivo changes in osteocyte protein expression when Ca2+ and actin dynamics are disrupted

Mechanical loading of osteocytes induces robust, intracellular calcium , Ca²⁺ oscillations and is also known to regulate osteocyte protein expression, including proteins capable of modulating osteoblast and osteoclast activity, cells that respectively build and resorb bone. Despite these clear and important roles in the adaptive process, to date, no studies have directly connected Ca²⁺-mediated mechanosensitivity to protein changes in osteocytes and few have linked Ca²⁺ oscillations to long-term adaptive responses.

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