Bone canonical Wnt signaling is downregulated in type 2 diabetes and associates with higher advanced glycation end-products (AGEs) content and reduced bone strength.

Type 2 diabetes (T2D) is associated with higher fracture risk, despite normal or high bone mineral density. We reported that bone formation genes (SOST and RUNX2) and advanced glycation end-products (AGEs) were impaired in T2D. We investigated Wnt signaling regulation and its association with AGEs accumulation and bone strength in T2D from bone tissue of 15 T2D and 21 non-diabetic postmenopausal women undergoing hip arthroplasty. Bone histomorphometry revealed a trend of low mineralized volume in T2D (T2D 0.249% [0.156-0.366]) vs non-diabetic subjects 0.352% [0.269-0.454]; p=0.053, as well as reduced bone strength (T2D 21.60 MPa [13.46-30.10] vs non-diabetic subjects 76.24 MPa [26.81-132.9]; p=0.002). We also showed that gene expression of Wnt agonists LEF-1 (p=0.0136) and WNT10B (p=0.0302) were lower in T2D. Conversely, gene expression of WNT5A (p=0.0232), SOST (p<0.0001), and GSK3B (p=0.0456) were higher, while collagen (COL1A1) was lower in T2D (p=0.0482). AGEs content was associated with SOST and WNT5A (r=0.9231, p<0.0001; r=0.6751, p=0.0322), but inversely correlated with LEF-1 and COL1A1 (r=-0.7500, p=0.0255; r=-0.9762, p=0.0004). SOST was associated with glycemic control and disease duration (r=0.4846, p=0.0043; r=0.7107, p=0.00174), whereas WNT5A and GSK3B were only correlated with glycemic control (r=0.5589, p=0.0037; r=0.4901, p=0.0051). Finally, Young's modulus was negatively correlated with SOST (r=-0.5675, p=0.0011), AXIN2 (r=-0.5523, p=0.0042), and SFRP5 (r=-0.4442, p=0.0437), while positively correlated with LEF-1 (r=0.4116, p=0.0295) and WNT10B (r=0.6697, p=0.0001). These findings suggest that Wnt signaling and AGEs could be the main determinants of bone fragility in T2D.

Type 2 diabetes is a long-term metabolic disease characterised by chronic high blood sugar levels. This in turn has a negative impact on the health of other tissues and organs, including bones. Type 2 diabetes patients have an increased risk of fracturing bones compared to non-diabetics. This is particularly true for fragility fractures, which are fractures caused by falls from a short height (i.e., standing height or less), often affecting hips or wrists. Usually, a lower bone density is associated with higher risk of fractures. However, patients with type 2 diabetes have increased bone fragility despite normal or higher bone density. One reason for this could be the chronically high levels of blood sugar in type 2 diabetes, which alter the properties of proteins in the body. It has been shown that the excess sugar molecules effectively 'react' with many different proteins, producing harmful compounds in the process, called Advanced Glycation End-products, or AGEs. AGEs are ­ in turn ­thought to affect the structure of collagen proteins, which help hold our tissues together and decrease bone strength. However, the signalling pathways underlying this process are still unclear. To find out more, Leanza et al. studied a signalling molecule, called sclerostin, which inhibits a signalling pathway that regulates bone formation, known as Wnt signaling. The researchers compared bone samples from both diabetic and non-diabetic patients, who had undergone hip replacement surgery. Analyses of the samples, using a technique called real-time-PCR, revealed that gene expression of sclerostin was increased in samples of type 2 diabetes patients, which led to a downregulation of Wnt signaling related genes. Moreover, the downregulation of Wnt genes was correlated with lower bone strength (which was measured by compressing the bone tissue). Further biochemical analysis of the samples revealed that higher sclerostin activity was also associated with higher levels of AGEs. These results provide a clearer understanding of the biological mechanisms behind compromised bone strength in diabetes. In the future, Leanza et al. hope that this knowledge will help us develop treatments to reduce the risk of bone complications for type 2 diabetes patients.

Assessment of bovine cortical bone fracture behavior using impact microindentation as a surrogate of fracture toughness.

The fracture behavior of bone is critically important for evaluating its mechanical competence and ability to resist fractures. Fracture toughness is an intrinsic material property that quantifies a material's ability to withstand crack propagation under controlled conditions. However, properly conducting fracture toughness testing requires the access to calibrated mechanical load frames and the destructive testing of bone samples, and therefore fracture toughness tests are clinically impractical. Impact microindentation mimicks certain aspects of fracture toughness measurements, but its relationship with fracture toughness remains unknown. In this study, we aimed to compare measurements of notched fracture toughness and impact microindentation in fresh and boiled bovine bone. Skeletally mature bovine bone specimens (n = 48) were prepared, and half of them were boiled to denature the organic matrix, while the other half remained preserved in frozen conditions. All samples underwent a notched fracture toughness test to determine their resistance to crack initiation (KIC) and an impact microindentation test using the OsteoProbe to obtain the Bone Material Strength index (BMSi). Boiling the bone samples increased the denatured collagen content, while mineral density and porosity remained unaffected. The boiled bones also showed significant reduction in both KIC (P < .0001) and the average BMSi (P < .0001), leading to impaired resistance of bone to crack propagation. Remarkably, the average BMSi exhibited a high correlation with KIC (r = 0.86; P < .001). A ranked order difference analysis confirmed the excellent agreement between the 2 measures. This study provides the first evidence that impact microindentation could serve as a surrogate measure for bone fracture behavior. The potential of impact microindentation to assess bone fracture resistance with minimal sample disruption could offer valuable insights into bone health without the need for cumbersome testing equipment and sample destruction.

Analysis of Infiltrating Immune Cells Following Intervertebral Disc Injury Reveals Recruitment of Gamma-Delta (γδ) T cells in Female Mice.

Inadequate repair of injured intervertebral discs (IVD) leads to degeneration and contributes to low back pain. Infiltrating immune cells into damaged musculoskeletal tissues are critical mediators of repair, yet little is known about their identities, roles, and temporal regulation following IVD injury. By analyzing longitudinal changes in gene expression, tissue morphology, and the dynamics of infiltrating immune cells following injury, we characterize sex-specific differences in immune cell populations and identify the involvement of previously unreported immune cell types, γδ and NKT cells. Cd3+Cd4-Cd8- T cells are the largest infiltrating lymphocyte population with injury, and we identified the presence of γδ T cells in this population in female mice specifically, and NKT cells in males. Injury-mediated IVD degeneration was prevalent in both sexes, but more severe in males. Sex-specific degeneration may be associated with the differential immune response since γδ T cells have potent anti-inflammatory roles and may mediate IVD repair.

Aging impairs the osteocytic regulation of collagen integrity and bone quality.

Poor bone quality is a major factor in skeletal fragility in elderly individuals. The molecular mechanisms that establish and maintain bone quality, independent of bone mass, are unknown but are thought to be primarily determined by osteocytes. We hypothesize that the age-related decline in bone quality results from the suppression of osteocyte perilacunar/canalicular remodeling (PLR), which maintains bone material properties. We examined bones from young and aged mice with osteocyte-intrinsic repression of TGFß signaling (TßRIIocy-/-) that suppresses PLR. The control aged bone displayed decreased TGFß signaling and PLR, but aging did not worsen the existing PLR suppression in male TßRIIocy-/- bone. This relationship impacted the behavior of collagen material at the nanoscale and tissue scale in macromechanical tests. The effects of age on bone mass, density, and mineral material behavior were independent of osteocytic TGFß. We determined that the decline in bone quality with age arises from the loss of osteocyte function and the loss of TGFß-dependent maintenance of collagen integrity.

Multi-omics resources for the Australian southern stuttering frog (Mixophyes australis) reveal assorted antimicrobial peptides.

The number of genome-level resources for non-model species continues to rapidly expand. However, frog species remain underrepresented, with up to 90% of frog genera having no genomic or transcriptomic data. Here, we assemble the first genomic and transcriptomic resources for the recently described southern stuttering frog (Mixophyes australis). The southern stuttering frog is ground-dwelling, inhabiting naturally vegetated riverbanks in south-eastern Australia. Using PacBio HiFi long-read sequencing and Hi-C scaffolding, we generated a high-quality genome assembly, with a scaffold N50 of 369.3 Mb and 95.1% of the genome contained in twelve scaffolds. Using this assembly, we identified the mitochondrial genome, and assembled six tissue-specific transcriptomes. We also bioinformatically characterised novel sequences of two families of antimicrobial peptides (AMPs) in the southern stuttering frog, the cathelicidins and ß-defensins. While traditional peptidomic approaches to peptide discovery have typically identified one or two AMPs in a frog species from skin secretions, our bioinformatic approach discovered 12 cathelicidins and two ß-defensins that were expressed in a range of tissues. We investigated the novelty of the peptides and found diverse predicted activities. Our bioinformatic approach highlights the benefits of multi-omics resources in peptide discovery and contributes valuable genomic resources in an under-represented taxon.

TRPV4 differentially controls inflammatory cytokine networks during static and dynamic compression of the intervertebral disc.

Background: The ion channel transient receptor potential vanilloid 4 (TRPV4) critically transduces mechanical forces in the IVD, and its inhibition can prevent IVD degeneration due to static overloading. However, it remains unknown whether different modes of loading signals through TRPV4 to regulate the expression of inflammatory cytokines. We hypothesized that TRPV4 signaling is essential during static and dynamic loading to mediate homeostasis and mechanotransduction. Methods: Mouse functional spine units were isolated and either cyclically compressed for 5 days (1 Hz, 1 h, 10% strain) or statically compressed (24 h, 0.2 MPa). Conditioned media were monitored at 6 h, 24 h, 2 days, and 5 days, with and without TRPV4 inhibition. Effects of TRPV4 activation was also evaluated without loading. The media was analyzed for a panel of 44 cytokines using a microbead array and then a correlative network was constructed to explore the regulatory relationships during loading and TRPV4 inhibition. After the loading regimen, the IVDs were evaluated histologically for degeneration. Results: Activation of TRPV4 led to an increase interleukin-6 (IL-6) family of cytokines (IL-6, IL-11, IL-16, and leukemia inhibitory factor [LIF]) and decreased the T-cell (CCL3, CCL4, CCL17, CCL20, CCL22, and CXCL10) and monocyte (CCL2 and CCL12) recruiting chemokines by the IVD. Dynamic and static loading each provoked unique chemokine correlation networks. The inhibition of TRPV4 during dynamic loading dysregulated the relationship between LIF and other cytokines, while the inhibition of TRPV4 during static loading disrupted the connectivity of IL-16 and VEGFA. Conclusions: We demonstrated that TRPV4 critically mediates the cytokine production following dynamic and static loading. The activation of TRPV4 upregulated a diverse set of cytokines that may suppress the chemotaxis of T-cells and monocytes, implicating the role of TRPV4 in maintaining the immune privilege of healthy IVD.

Preparing recombinant "Split AEP" for protein labeling.

A variant originated from Oldenlandia affinis asparaginyl ligase, OaAEP1-C247A, has emerged as an ideal tool for protein labeling. However, its preparation was laborious and time-consuming. It is recombinantly produced as a zymogen, requiring acid activation and four chromatographic steps; despite these extensive steps, the catalytically active enzyme exhibited only moderate purity. Here, we report a novel preparation protocol, in which the cap and catalytically active core domains are produced as separate entities. The active enzyme can be obtained in two chromatographic steps, immobilized metal affinity chromatography (IMAC) and size exclusion chromatography (SEC), with no acid activation required, thereby shortening the purification procedure from at least 2 days to less than 6 h. In addition to the original C247A mutation which enhanced reaction with various amino nucleophiles, an extra D29E mutation was introduced to prevent self-cleavage, which led to noticeable improvements in homogeneity and activity of the enzyme. Indeed, the resulting "split AEP" (i.e., core domain of OaAEP1-D29E/C247A) exhibited improved catalytic efficiency constant (kcat/KM) that was found to be ∼3-fold higher than that of the original acid-activated counterpart (OaAEP1-C247A). Furthermore, we described a protein labeling protocol that couples the enzymatic reaction with an irreversible chemical transformation, thereby enabling high conversion of labeled protein with a lowered amount of reagent. Precisely, an alternative Asn-Cys-Leu (NCL) recognition sequence was used for substrate recognition. As the byproduct contains an N-terminal cysteine, it can be transformed into an inert 1,2 aminothiol motif by reacting with formylphenyl boronic acid (FPBA). Finally, the opportunities and challenges associated with the use of asparaginyl ligase are discussed.

Comparing shades of darkness: trolling victims' experiences on social media vs. online gaming.

Although there is ample literature available on toxicity in games, as there is regarding trolling on social media, there are few to no cross-platform studies on toxicity and trolling. In other words, the extant literature focuses on one platform at a time instead of comparing and contrasting them. The present work aims to rectify this gap by analyzing interviews from a larger study of 22 self-proclaimed victims of in-game trolling to not only determine whether social media or gaming communities are considered more toxic but also to explore how definitions of the word 'trolling' change depending on the platform in question. We found that while definitions of in-game trolling behavior focused on behavioral styles of trolling (e.g., throwing one's avatar into enemy fire to disadvantage one's team, and blocking other players' avatars' movement), social media trolling is defined by more sinister actions such as misinformation spreading and 'canceling' other users. We also found that gaming is perceived as generally more toxic than social media, often due to company policies or lack thereof. Practical and theoretical implications for the study of toxicity in all online communities - gaming or social-media based - are discussed.

Foot Plantar Pressure Profile Alteration after Microsurgical Great Toe-to-thumb Transfer.

Background: Microsurgical great toe-to-thumb transfer (mGTT) is a widely used procedure when immediate replantation of thumb is not feasible. The aim of this study was to investigate the alteration of plantar pressure profile of the donor foot after mGTT. Methods: Twenty patients receiving microsurgical great toe-to-hand transfer between 1985 to 2014, and 16 healthy subjects were recruited. Group 1 consisted of 20 feet receiving mGTT, whereas group 2 consisted of 32 normal feet as control. The flap design in this study was to preserve 1 cm of the proximal phalanx to maintain the attachment of the plantar aponeurosis and intrinsic muscles. The Taiwan Chinese version of the Foot Function Index was used for patient-reported outcome measurement. A novel Emed-X system was used for dynamic plantar pressure measurement. A total of four parameters were collected, including peak pressure, contact area, contact time, and pressure-time integral. Results: In group 1, the peak pressure redistributed under the first metatarsal bone and was significantly higher than group 2 (P < 0.05). There was no significant change of the contact area between the midfoot region of group 1 and group 2 (P > 0.05). Furthermore, similar foot clearance efficiency was demonstrated in group 1 and group 2 (P > 0.05). Conclusions: The windlass effect of the foot will not be affected when performing mGTT with preservation of 1 cm of the proximal phalanx. Therefore, this surgical procedure is highly recommended for clinical application.

Assessment of bovine cortical bone fracture behavior using impact microindentation as a surrogate of fracture toughness.

The fracture behavior of bone is critically important for assessing its mechanical competence and ability to resist fractures. Fracture toughness, which quantifies a material's resistance to crack propagation under controlled geometry, is regarded as the gold standard for evaluating a material's resistance to fracture. However properly conducting this test requires access to calibrated mechanical load frames the destruction of the bone samples, making it impractical for obtaining clinical measurement of bone fracture. Impact microindentation offers a potential alternative by mimicking certain aspects of fracture toughness measurements, but its relationship with mechanistic fracture toughness remains unknown. In this study, we aimed to compare measurements of notched fracture toughness and impact microindentation in fresh and boiled bovine bone. Skeletally mature bovine bone specimens (n=48) were prepared, and half of them were boiled to denature the organic matrix, while the other half remained preserved in frozen conditions. Notched fracture toughness tests were conducted on all samples to determine Initiation toughness (KIC), and an impact microindentation test using the OsteoProbe was performed to obtain the Bone Material Strength index. Boiling the bone samples resulted increased the denatured collagen without affecting mineral density or porosity. The boiled bones also showed significant reduction in both KIC (p < 0.0001) and the average Bone Material Strength index (p < 0.0001), leading to impaired resistance of bone to crack propagation. Remarkably, the average Bone Material Strength index exhibited a high correlation with KIC (r = 0.86; p < 0.001). The ranked order difference analysis confirmed excellent agreement between the two measures. This study provides the first evidence that impact microindentation could serve as a surrogate measure for bone fracture behavior. The potential of impact microindentation to non-destructively assess bone fracture resistance could offer valuable insights into bone health without the need for elaborate testing equipment and sample destruction.

Relative Effects of Radiation-Induced Changes in Bone Mass, Structure, and Tissue Material on Vertebral Strength in a Rat Model.

The observed increased risk of fracture after cancer radiation therapy is presumably due to a radiation-induced reduction in whole-bone strength. However, the mechanisms for impaired strength remain unclear, as the increased fracture risk is not fully explained by changes in bone mass. To provide insight, a small animal model was used to determine how much of this whole-bone weakening effect for the spine is attributable to changes in bone mass, structure, and material properties of the bone tissue and their relative effects. Further, because women have a greater risk of fracture after radiation therapy than men, we investigated if sex had a significant influence on bone's response to irradiation. Fractionated in vivo irradiation (10 × 3 Gy) or sham irradiation (0 Gy) was administered daily to the lumbar spine in twenty-seven 17-week-old Sprague-Dawley rats (n = 6-7/sex/group). Twelve weeks after final treatment, animals were euthanized, and lumbar vertebrae (L4 and L5 ) were isolated. Using a combination of biomechanical testing, micro-CT-based finite element analysis, and statistical regression analysis, we separated out the effect of mass, structural, and tissue material changes on vertebral strength. Compared with the sham group (mean ± SD strength = 420 ± 88 N), the mean strength of the irradiated group was lower by 28% (117 N/420 N, p < 0.0001). Overall, the response of treatment did not differ with sex. By combining results from both general linear regression and finite element analyses, we calculated that mean changes in bone mass, structure, and material properties of the bone tissue accounted for 56% (66 N/117 N), 20% (23 N/117 N), and 24% (28 N/117 N), respectively, of the overall change in strength. As such, these results provide insight into why an elevated clinical fracture risk for patients undergoing radiation therapy is not well explained by changes in bone mass alone. © 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Bullying, harassment, and discrimination of musculoskeletal researchers and the impact of the COVID-19 pandemic: an international study.

PURPOSE: Bullying, harassment, and discrimination (BHD) are prevalent in academic, scientific, and clinical departments, particularly orthopedic surgery, and can have lasting effects on victims. As it is unclear how BHD affects musculoskeletal (MSK) researchers, the following study assessed BHD in the MSK research community and whether the COVID-19 pandemic, which caused hardships in other industries, had an impact. METHODS: A web-based anonymous survey was developed in English by ORS Spine Section members to assess the impact of COVID-19 on MSK researchers in North America, Europe, and Asia, which included questions to evaluate the personal experience of researchers regarding BHD. RESULTS: 116 MSK researchers completed the survey. Of respondents, 34.5% (n = 40) focused on spine, 30.2% (n = 35) had multiple areas of interest, and 35.3% (n = 41) represented other areas of MSK research. BHD was observed by 26.7% (n = 31) of respondents and personally experienced by 11.2% (n = 13), with mid-career faculty both observing and experiencing the most BHD. Most who experienced BHD (53.8%, n = 7) experienced multiple forms. 32.8% (n = 38) of respondents were not able to speak out about BHD without fear of repercussions, with 13.8% (n = 16) being unsure about this. Of those who observed BHD, 54.8% (n = 17) noted that the COVID-19 pandemic had no impact on their observations. CONCLUSIONS: To our knowledge, this is the first study to address the prevalence and determinants of BHD among MSK researchers. MSK researchers experienced and observed BHD, while many were not comfortable reporting and discussing violations to their institution. The COVID-19 pandemic had mixed-effects on BHD. Awareness and proactive policy changes may be warranted to reduce/eliminate the occurrence of BHD in this community.

embryoTox: Using Graph-Based Signatures to Predict the Teratogenicity of Small Molecules.

Teratogenic drugs can lead to extreme fetal malformation and consequently critically influence the fetus's health, yet the teratogenic risks associated with most approved drugs are unknown. Here, we propose a novel predictive tool, embryoTox, which utilizes a graph-based signature representation of the chemical structure of a small molecule to predict and classify molecules likely to be safe during pregnancy. embryoTox was trained and validated using in vitro bioactivity data of over 700 small molecules with characterized teratogenicity effects. Our final model achieved an area under the receiver operating characteristic curve (AUC) of up to 0.96 on 10-fold cross-validation and 0.82 on nonredundant blind tests, outperforming alternative approaches. We believe that our predictive tool will provide a practical resource for optimizing screening libraries to determine effective and safe molecules to use during pregnancy. To provide a simple and integrated platform to rapidly screen for potential safe molecules and their risk factors, we made embryoTox freely available online at https://biosig.lab.uq.edu.au/embryotox/.

Modulation of TRPV4 protects against degeneration induced by sustained loading and promotes matrix synthesis in the intervertebral disc.

While it is well known that mechanical signals can either promote or disrupt intervertebral disc (IVD) homeostasis, the molecular mechanisms for transducing mechanical stimuli are not fully understood. The transient receptor potential vanilloid 4 (TRPV4) ion channel activated in isolated IVD cells initiates extracellular matrix (ECM) gene expression, while TRPV4 ablation reduces cytokine production in response to circumferential stretching. However, the role of TRPV4 on ECM maintenance during tissue-level mechanical loading remains unknown. Using an organ culture model, we modulated TRPV4 function over both short- (hours) and long-term (days) and evaluated the IVDs' response. Activating TRPV4 with the agonist GSK101 resulted in a Ca2+ flux propagating across the cells within the IVD. Nuclear factor (NF)-κB signaling in the IVD peaked at 6 h following TRPV4 activation that subsequently resulted in higher interleukin (IL)-6 production at 7 days. These cellular responses were concomitant with the accumulation of glycosaminoglycans and increased hydration in the nucleus pulposus that culminated in higher stiffness of the IVD. Sustained compressive loading of the IVD resulted in elevated NF-κB activity, IL-6 and vascular endothelial growth factor A (VEGFA) production, and degenerative changes to the ECM. TRPV4 inhibition using GSK205 during loading mitigated the changes in inflammatory cytokines, protected against IVD degeneration, but could not prevent ECM disorganization due to mechanical damage in the annulus fibrosus. These results indicate TRPV4 plays an important role in both short- and long-term adaptations of the IVD to mechanical loading. The modulation of TRPV4 may be a possible therapeutic for preventing load-induced IVD degeneration.

Quantative MRI predicts tendon mechanical behavior, collagen composition, and organization.

Quantitative magnetic resonance imaging (qMRI) measures have provided insights into the composition, quality, and structure-function of musculoskeletal tissues. Low signal-to-noise ratio has limited application to tendon. Advances in scanning sequences and sample positioning have improved signal from tendon allowing for evaluation of structure and function. The purpose of this study was to elucidate relationships between tendon qMRI metrics (T1, T2, T1ρ and diffusion tensor imaging [DTI] metrics) with tendon tissue mechanics, collagen concentration and organization. Sixteen human Achilles tendon specimens were collected, imaged with qMRI, and subjected to mechanical testing with quantitative polarized light imaging. T2 values were related to tendon mechanics [peak stress (rsp = 0.51, p = 0.044), equilibrium stress (rsp = 0.54, p = 0.033), percent relaxation (rsp = -0.55, p = 0.027), hysteresis (rsp = -0.64, p = 0.007), linear modulus (rsp = 0.67, p = 0.009)]. T1ρ had a statistically significant relationship with percent relaxation (r = 0.50, p = 0.048). Collagen content was significantly related to DTI measures (range of r = 0.56-0.62). T2 values from a single slice of the midportion of human Achilles tendons were strongest predictors of tendon tensile mechanical metrics. DTI diffusivity indices (mean diffusivity, axial diffusivity, radial diffusivity) were strongly correlated with collagen content. These findings build on a growing body of literature supporting the feasibility of qMRI to characterize tendon tissue and noninvasively measure tendon structure and function. Statement of Clinical Significance: Quantitative MRI can be applied to characterize tendon tissue and is a noninvasive measure that relates to tendon composition and mechanical behavior.

Geodesic Logistic Analysis of Lumbar Spine Intervertebral Disc Shapes in Supine and Standing Positions.

Non-specific lower back pain (LBP) is a world-wide public health problem that affects people of all ages. Despite the high prevalence of non-specific LBP and the associated economic burdens, the pathoanatomical mechanisms for the development and course of the condition remain unclear. While intervertebral disc degeneration (IDD) is associated with LBP, there is overlapping occurrence of IDD in symptomatic and asymptomatic individuals, suggesting that degeneration alone cannot identify LBP populations. Previous work has been done trying to relate linear measurements of compression obtained from Magnetic Resonance Imaging (MRI) to pain unsuccessfully. To bridge this gap, we propose to use advanced non-Euclidean statistical shape analysis methods to develop biomarkers that can help identify symptomatic and asymptomatic adults who might be susceptible to standing-induced LBP. We scanned 4 male and 7 female participants who exhibited lower back pain after prolonged standing using an Open Upright MRI. Supine and standing MRIs were obtained for each participant. Patients reported their pain intensity every fifteen minutes within a period of 2 h. Using our proposed geodesic logistic regression, we related the structure of their lower spine to pain and computed a regression model that can delineate lower spine structures using reported pain intensities. These results indicate the feasibility of identifying individuals who may suffer from lower back pain solely based on their spinal anatomy. Our proposed spinal shape analysis methodology have the potential to provide powerful information to the clinicians so they can make better treatment decisions.

Single Cell RNA-Sequence Analyses Reveal Uniquely Expressed Genes and Heterogeneous Immune Cell Involvement in the Rat Model of Intervertebral Disc Degeneration.

Intervertebral disc (IVD) degeneration is characterized by a loss of cellularity, and changes in cell-mediated activity that drives anatomic changes to IVD structure. In this study, we used single-cell RNA-sequencing analysis of degenerating tissues of the rat IVD following lumbar disc puncture. Two control, uninjured IVDs (L2-3, L3-4) and two degenerated, injured IVDs (L4-5, L5-6) from each animal were examined either at the two- or eight-week post-operative time points. The cells from these IVDs were extracted and transcriptionally profiled at the single-cell resolution. Unsupervised cluster analysis revealed the presence of four known cell types in both non-degenerative and degenerated IVDs based on previously established gene markers: IVD cells, endothelial cells, myeloid cells, and lymphoid cells. As a majority of cells were associated with the IVD cell cluster, sub-clustering was used to further identify the cell populations of the nucleus pulposus, inner and outer annulus fibrosus. The most notable difference between control and degenerated IVDs was the increase of myeloid and lymphoid cells in degenerated samples at two- and eight-weeks post-surgery. Differential gene expression analysis revealed multiple distinct cell types from the myeloid and lymphoid lineages, most notably macrophages and B lymphocytes, and demonstrated a high degree of immune specificity during degeneration. In addition to the heterogenous infiltrating immune cell populations in the degenerating IVD, the increased number of cells in the AF sub-cluster expressing Ngf and Ngfr, encoding for p75NTR, suggest that NGF signaling may be one of the key mediators of the IVD crosstalk between immune and neuronal cell populations. These findings provide the basis for future work to understand the involvement of select subsets of non-resident cells in IVD degeneration.

Contrast-enhanced microCT evaluation of degeneration following partial and full width injuries to the mouse lumbar intervertebral disc.

A targeted injury to the mouse intervertebral disc (IVD) is often used to recapitulate the degenerative cascade of the human pathology. Since injuries can vary in magnitude and localization, it is critical to examine the effects of different injuries on IVD degeneration. We thus evaluated the degenerative progression resulting from either a partial- or full-width injury to the mouse lumbar IVD using contrast-enhanced micro-computed tomography and histological analyses. A lateral-retroperitoneal surgical approach was used to access the lumbar IVD, and the injuries to the IVD were produced by either incising one side of the annulus fibrosus or puncturing both sides of the annulus fibrosus. Female C57BL/6J mice of 3-4 months age were used in this study. They were divided into three groups to undergo partial-width, full-width, or sham injuries. The L5/6 and L6/S1 lumbar IVDs were surgically exposed, and then the L6/S1 IVDs were injured using either a surgical scalpel (partial-width) or a 33G needle (full-width), with the L5/6 serving as an internal control. These animals recovered and then euthanized at either 2-, 4-, or 8-weeks after surgery for evaluation. The IVDs were assessed for degeneration using contrast-enhanced microCT (CEµCT) and histological analysis. The high-resolution 3D CEµCT evaluation of the IVD confirmed that the respective injuries were localized within one side of the annulus fibrosus or spanned the full width of the IVD. The full-width injury caused significant deteriorations in the nucleus pulposus, annulus fibrous and at the interfaces after 2 weeks, which was sustained through the 8 weeks, while the partial width injury caused localized disruptions that remained limited to the annulus fibrosus. The use of CEµCT revealed distinct IVD degeneration profiles resulting from partial- and full-width injuries. The partial width injury may serve as an alternative model for IVD degeneration resulting from localized annulus fibrosus injuries.

Rates of MRI simulator utilisation in a tertiary cancer therapy centre.

Magnetic resonance imaging (MRI) is increasingly being integrated into the radiation oncology workflow, due to its improved soft tissue contrast without additional exposure to ionising radiation. A review of MRI utilisation according to evidence based departmental guidelines was performed. Guideline utilisation rates were calculated to be 50% (true utilisation rate was 46%) of all new cancer patients treated with adjuvant or curative intent, excluding simple skin and breast cancer patients. Guideline utilisation rates were highest in the lower gastrointestinal and gynaecological subsites, with the lowest being in the upper gastrointestinal and thorax subsites. Head and neck (38% vs 45%) and CNS (46% vs 67%) cancers had the largest discrepancy between true and guideline utilisation rates due to unnamed reasons and non-contemporaneous diagnostic imaging respectively. This report outlines approximate MRI utilisation rates in a tertiary radiation oncology service and may help guide planning for future departments contemplating installation of an MRI simulator.

The degenerative impact of hyperglycemia on the structure and mechanics of developing murine intervertebral discs.

Introduction: Diabetes has long been implicated as a major risk factor for intervertebral disc (IVD) degeneration, interfering with molecular signaling and matrix biochemistry, which ultimately aggravates the progression of the disease. Glucose content has been previously shown to influence structural and compositional changes in engineered discs in vitro, impeding fiber formation and mechanical stability. Methods: In this study, we investigated the impact of diabetic hyperglycemia on young IVDs by assessing biochemical composition, collagen fiber architecture, and mechanical behavior of discs harvested from 3- to 4-month-old db/db mouse caudal spines. Results: We found that discs taken from diabetic mice with elevated blood glucose levels demonstrated an increase in total glycosaminoglycan and collagen content, but comparable advanced glycation end products (AGE) levels to wild-type discs. Diabetic discs also contained ill-defined boundaries between the nucleus pulposus and annulus fibrosus, with the latter showing a disorganized and unaligned collagen fiber network at this same boundary. Conclusions: These compositional and structural changes had a detrimental effect on function, as the diabetic discs were twice as stiff as their wild-type counterparts and demonstrated a significant resistance to deformation. These results indicate that diabetes may predispose the young disc to DDD later in life by altering patterns of extracellular matrix deposition, fiber formation, and motion segment mechanics independently of AGE accumulation.