Biology subjects

Tomlinson, R. E.

Publications and source records attributed to Tomlinson, R. E..

9 recordsLinked to original sources

bioRxiv2025

COX2-independent and COX2-dependent effects of naproxen on bone quality, osteocytes, and fatigue fracture healing in male and female mice

Regular non-steroidal anti-inflammatory drug (NSAID) use increases stress fracture risk, but the mechanisms remain unclear. Here, we used Ptgs2-Y385F mice, which lack cyclooxygenase 2 (COX2) enzyme activity, to test the hypothesis that naproxen decreases strain adaptive bone remodeling in a COX2-dependent manner and decreases bone toughness and fracture resistance through COX2-independent effects. MicroCT and mechanical testing showed minimal baseline differences between Ptgs2-Y385F and wild-type (WT) mice. Following non-damaging forelimb compression, naproxen decreased load-induced bone formation in WT, but not Ptgs2-Y385F mice, consistent with a COX2-dependent effect. In contrast, naproxen reduced bone toughness and post-yield deformation across genotypes and doses, supporting a COX2-independent mechanism. Histologically, naproxen increased empty osteocyte lacunae in both genotypes, while osteoblast number, total lacunae, and perilacunar labeling were increased only in WT mice. Naproxen pre-treatment before fatigue fracture produced mild, sex-specific effects on fracture initiation and healing. Analysis of osteocyte dendritic networks in uninjured femurs revealed sexually dimorphic changes in dendrite number and density in naproxen-treated mice as compared to vehicle. In total, naproxen influences bone through both COX2-dependent and COX2-independent mechanisms, with some sexual dimorphism. These findings confirm that regular NSAID usage compromises skeletal health and underscore the need for new pain management strategies.

bioengineering↗

bioRxiv2025

The Effect of Celecoxib and muMab911 on Strain Adaptive Bone Remodeling and Fracture Repair in Female Mice: Implications for Rapidly Progressive Osteoarthritis

Debilitating pain is the primary clinical feature of osteoarthritis (OA) that drives the enormous healthcare costs. Osteoarthritis-related pain is often treated with non-steroidal anti-inflammatory drugs (NSAIDs), which effectively relieve pain and inflammation by inhibition of prostaglandin synthesis. Antibodies directed against nerve growth factor (NGF) were tested some time ago as an alternative potential analgesic for musculoskeletal pain, including osteoarthritis-related pain. Unfortunately, clinical development of these drugs was put on hold due to adverse outcomes - primarily rapidly progressive osteoarthritis. Both prostaglandin synthesis and NGF have been implicated as critical mediators of strain adaptive bone remodeling, which may play a role in rapid osteoarthritis progression. Therefore, this study was designed to investigate the effects of celecoxib, an NSAID, and muMab911, an anti-NGF antibody, as well as the combination therapy on strain adaptive bone remodeling, bone mass and geometry, and bone healing in a murine model. Adult female C57BL/6J mice received celecoxib through drinking water, up to 3 IP injections of muMab911, or both treatments over a period of two weeks. As expected, all treatments were effective for relieving injury-associated pain. Consistent with previous studies, we found that celecoxib alone and in combination with muMab911 impaired periosteal load-induced bone formation induced by axial forelimb compression. Furthermore, both treatments had minimal effects on osteoblast and osteocyte populations, bone structural and material properties, and cortical and trabecular bone mass. Mice subjected to damaging axial forelimb compression did not have impaired healing or callus morphology due to treatment. Nonetheless, both drugs decreased NGF expression during fracture healing. In total, our results indicate that these medications both suppress NGF expression and load-induced bone formation which may support rapid OA disease progression by diminishing strain adaptive bone remodeling in subchondral bone of OA patients.

bioengineering↗

bioRxiv2022

Toll-like receptor 4 signaling in osteoblasts is required for load-induced bone formation in mice

During skeletal development, expression of nerve growth factor (NGF) leads to the survival of afferent sensory nerves that express neurotrophic tyrosine kinase receptor type 1 (TrkA), the high affinity receptor for NGF. In adulthood, NGF is expressed by mature osteoblasts following mechanical loading and signals through TrkA receptors in resident sensory nerves to support load-induced bone formation. However, the regulation of NGF in osteoblasts following loading is not well understood. In this study, we sought to determine whether osteoblastic expression of toll-like receptor 4 (TLR4), a key receptor in the NF-{kappa}B signaling pathway, is required to initiate NGF-TrkA signaling to support skeletal adaptation following mechanical loading. First, we observed that NF-{kappa}B inhibition reduces NGF expression induced by axial forelimb compression. Moreover, we observed that TLR4+ periosteal cells are increased after mechanical loading. Therefore, we generated a novel mouse model in which Tlr4 is selectively removed from the mature osteoblast lineage. Although Tlr4 conditional knockout mice have normal skeletal mass and strength in adulthood, the loss of TLR4 signaling results in significant reductions in periosteal lamellar bone formation following axial forelimb compression. Furthermore, we demonstrated that the upregulation of Ngf following application of fluid shear stress to calvarial osteoblasts is significantly reduced by NF-{kappa}B and TLR4 inhibitors. Finally, RNA sequencing demonstrated that the deficits in load-induced bone formation in CKO mice can be attributed to dysregulated inflammatory signaling. In total, our study reveals a novel role for TLR4 in skeletal adaptation to mechanical loading in bone, which may enable new therapeutic strategies for diseases of low bone mass and provide new targets for musculoskeletal pain relief.

cell biology↗

bioRxiv2022

Approximating Scaffold Printability Utilizing Computational Methods

Bioprinting facilitates the generation of complex, three-dimensional (3D), cell-based constructs for various applications. Although multiple bioprinting technologies have been developed, extrusion-based systems have become the dominant technology due to the diversity of materials (bioinks) that can be utilized, either individually or in combination. However, each bioink has unique material properties and extrusion characteristics that affect bioprinting utility, accuracy, and precision. Here, we have extended our previous work to achieve high precision (i.e., repeatability) across samples by optimizing bioink-specific printing parameters. Specifically, we hypothesized that an adaptive neuro-fuzzy inference system (ANFIS) could be used as a computational method to address the imprecision in 3D bioprinting test data and uncover the optimal printing parameters for a specific bioink that result in high accuracy and precision. To test this hypothesis, we have implemented an ANFIS model consisting of four inputs (bioink concentration, printing pressure, speed, and temperature) and a single output to quantify the precision (scaffold bioprinted linewidth range). We validate our use of the bioprinting precision index (BPI) with both standard and normalized printability factors. In total, our results indicate that computational methods are a cost-efficient measure to improve the precision and robustness of extrusion 3D bioprinting with gelatin-based bioinks.

bioengineering↗

bioRxiv2021

Comparison of Type-1 and Type-2 Fuzzy Systems forMineralization of Bioprinted Bone

Bioprinting is an emerging tissue engineering method used to generate cell-laden scaffolds with high spatial resolution. Bioprinting parameters, such as pressure, nozzle size, and speed, have a large influence on the quality of the bioprinted construct. Moreover, cell suspension density, cell culture period, and other critical biological parameters directly impact the biological function of the final product. Therefore, an approximation model that can be used to find the values of bioprinting parameters that will result in optimal bioprinted constructs is highly desired. Here, we propose type-1 and type-2 fuzzy systems to handle the uncertainty and imprecision in optimizing the input values. Specifically, we focus on the biological parameters, such as culture period, that can be used to maximize the output value (mineralization volume). To achieve a more accurate approximation, we have compared a type-2 fuzzy system with a type-1 fuzzy system using two levels of uncertainty. We hypothesized that type-2 fuzzy systems may be preferred in biological systems, due to the inherent vagueness and imprecision of the input data. Here, our results demonstrate that the type-2 fuzzy system with a high uncertainty boundary (30%) is superior to type-1 and type-2 with low uncertainty boundary fuzzy systems in the overall output approximation error for bone bioprinting inputs.

bioinformatics↗

bioRxiv2020

Utilizing Q-Learning to Generate 3D Vascular Networks for Bioprinting Bone

Bioprinting is an emerging tissue engineering method used to generate cell-laden scaffolds with high spatial resolution. Bioprinted vascularized bone grafts are a potential application of this technology that would meet a critical clinical need, since current approaches to volumetric bone repair have significant limitations. However, generation of vascular networks suitable for bioprinting is challenging. Here, we propose a novel Q-learning approach to quickly generate 3D vascular networks within patient-specific bone geometry that are optimized for bioprinting. First, the inlet and outlet locations are specified and the scenario is modeled using a grid world for initial agent training. Next, the path planned in the grid world environment is converted to a Bezier curve, which is then used to generate the final 3D vascularized bone model. The vessels generated using this procedure have minimal tortuosity, which increases the likelihood of successful bioprinting. Furthermore, the ability to specify inlet and outlet position is necessary for both surgical feasibility as well as generation of more complex vascular networks. In total, this study demonstrates the reliability of our reinforcement learning method for automated generation of 3D vascular networks within patient-specific geometry that can be used for bioprinting vascularized bone grafts.

bioengineering↗

bioRxiv2020

The TrkA agonist gambogic amide augments skeletal adaptation to mechanical loading through actions on sensory nerves and osteoblasts

The periosteal and endosteal surfaces of mature bone are densely innervated by sensory nerves expressing TrkA, the high-affinity receptor for nerve growth factor (NGF). In previous work, we demonstrated that administration of exogenous NGF significantly increased load-induced bone formation through the activation of Wnt signaling. However, the translational potential of NGF is limited by the induction of substantial mechanical and thermal hyperalgesia in mice and humans. Here, we tested the effect of gambogic amide (GA), a recently identified robust small molecule agonist for TrkA, on hyperalgesia and load-induced bone formation. Behavioral analysis was used to assess pain up to one week after axial forelimb compression. Contrary to our expectations, GA treatment was not associated with diminished use of the loaded forelimb or sensitivity to thermal stimulus. Furthermore, dynamic histomorphometry revealed a significant increase in relative periosteal bone formation rate as compared to vehicle treatment. Additionally, we found that GA treatment was associated with an increase in the number of osteoblasts per bone surface in loaded limbs as well as a significant upregulation of Wnt1, Wnt7b, and Ngf in loaded bones. To determine if these effects were exclusively mediated by NGF-TrkA signaling in sensory nerves, we cultured MC3T3-E1 cells for 7 or 14 days in osteogenic differentiation media containing NGF (50 ng/mL), GA (5, 50, or 500 nM), or vehicle (DMSO). After 7 days of culture, we observed increases in osteoblastic differentiation markers Runx2, Bglap2, and Sp7 in response to GA, whereas treatment with NGF was not different than vehicle. Only cells treated with the highest dose of GA (500 nM) had significantly impaired cell proliferation. In conclusion, our study indicates GA may be useful for augmenting skeletal adaptation to mechanical forces without inducing hyperalgesia through actions on both sensory nerves and osteoblasts.

bioengineering↗

bioRxiv2019

Ankylosis homologue (ANKH) controls extracellular citrate and pyrophosphate homeostasis and affects bone mechanical performance

The membrane protein Ankylosis homologue (ANKH, mouse orthologue: ANK) prevents mineralization of joint-space and articular cartilage. The accepted view is that ANKH mediates cellular release of inorganic pyrophosphate (PPi), a strong physiological inhibitor of mineralization. Using global metabolite profiling, we identified citrate as the most prominent metabolite leaving HEK293 cells in an ANKH-dependent manner. Although PPi levels were increased in culture medium of HEK293-ANKH cells, PPi was formed extracellularly after release of ATP and other nucleoside triphosphates. Ankank/ank mice, which lack functional ANK, had substantially reduced concentrations of citrate in plasma and urine, while citrate was undetectable in urine of a human patient lacking functional ANKH. Bone hydroxyapatite of Ankank/ank mice also contained markedly reduced levels of citrate and PPi and displayed diminished strength. Together, our data show that ANKH is a crucial factor in extracellular citrate and PPi homeostasis that is essential for normal bone development.

developmental biology↗

bioRxiv2018

First Whole-Body Three-Dimensional Tomographic Imaging of Alpha Particle Emitting Radium-223

Objective: Dose optimization and pharmacokinetic evaluation of alpha emitting Radium-223 dichloride (223RaCl2) by planar gamma camera or single photon emission computed tomographic (SPECT) imaging are hampered by the low photon abundance and injection activities. Here, we demonstrate SPECT of 223Ra using phantoms and small animal in vivo models. Methods: Line phantoms and mice bearing 223Ra were imaged using a next generation dedicated small animal SPECT by detecting the low energy photon emissions from 223Ra. Localization of the therapeutic agent was verified by whole body and whole limb autoradiography and its effect determined by immunofluorescence. Results: A state-of-the-art commercial small animal SPECT system equipped with a highly sensitive collimator enables collection of sufficient counts for three-dimensional reconstruction. Line sources of 223Ra in both air and in a water scattering phantom gave linear response functions with provide full-width-at-half-maximum of 1.45 mm. Early and late phase imaging of the pharmacokinetics of the radiopharmaceutical were captured. Uptake at sites of active bone remodeling were correlated with DNA damage from the alpha particle emissions. Conclusions: This work demonstrates the capability to noninvasively define the distribution of 223Ra, a recently approved alpha emitting radionuclide. This approach allows quantitative assessment of 223Ra distribution and may provide radiation dose optimization strategies to improve therapeutic response and ultimately to enable personalized treatment planning.

pharmacology and toxicology↗