TACAN Is an Ion Channel Involved in Sensing Mechanical Pain.

Mechanotransduction, the conversion of mechanical stimuli into electrical signals, is a fundamental process underlying essential physiological functions such as touch and pain sensing, hearing, and proprioception. Although the mechanisms for some of these functions have been identified, the molecules essential to the sense of pain have remained elusive. Here we report identification of TACAN (Tmem120A), an ion channel involved in sensing mechanical pain. TACAN is expressed in a subset of nociceptors, and its heterologous expression increases mechanically evoked currents in cell lines. Purification and reconstitution of TACAN in synthetic lipids generates a functional ion channel. Finally, a nociceptor-specific inducible knockout of TACAN decreases the mechanosensitivity of nociceptors and reduces behavioral responses to painful mechanical stimuli but not to thermal or touch stimuli. We propose that TACAN is an ion channel that contributes to sensing mechanical pain.

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.

Regional and age-related diversity of human mature oligodendrocytes.

Morphological and emerging molecular studies have provided evidence for heterogeneity within the oligodendrocyte population. To address the regional and age-related heterogeneity of human mature oligodendrocytes (MOLs) we applied single-cell RNA sequencing to cells isolated from cortical/subcortical, subventricular zone brain tissue samples, and thoracolumbar spinal cord samples. Unsupervised clustering of cells identified transcriptionally distinct MOL subpopulations across regions. Spinal cord MOLs, but not microglia, exhibited cell-type-specific upregulation of immune-related markers compared to the other adult regions. SVZ MOLs showed an upregulation of select number of development-linked transcription factors compared to other regions; however, pseudotime trajectory analyses did not identify a global developmental difference. Age-related analysis of cortical/subcortical samples indicated that pediatric MOLs, especially from under age 5, retain higher expression of genes linked to development and to immune activity with pseudotime analysis favoring a distinct developmental stage. Our regional and age-related studies indicate heterogeneity of MOL populations in the human CNS that may reflect developmental and environmental influences.

Single-Cell RNA-Seq Analysis of Cells from Degenerating and Non-Degenerating Intervertebral Discs from the Same Individual Reveals New Biomarkers for Intervertebral Disc Degeneration.

In this study, we used single-cell transcriptomic analysis to identify new specific biomarkers for nucleus pulposus (NP) and inner annulus fibrosis (iAF) cells, and to define cell populations within non-degenerating (nD) and degenerating (D) human intervertebral discs (IVD) of the same individual. Cluster analysis based on differential gene expression delineated 14 cell clusters. Gene expression profiles at single-cell resolution revealed the potential functional differences linked to degeneration, and among NP and iAF subpopulations. GO and KEGG analyses discovered molecular functions, biological processes, and transcription factors linked to cell type and degeneration state. We propose two lists of biomarkers, one as specific cell type, including C2orf40, MGP, MSMP, CD44, EIF1, LGALS1, RGCC, EPYC, HILPDA, ACAN, MT1F, CHI3L1, ID1, ID3 and TMED2. The second list proposes predictive IVD degeneration genes, including MT1G, SPP1, HMGA1, FN1, FBXO2, SPARC, VIM, CTGF, MGST1, TAF1D, CAPS, SPTSSB, S100A1, CHI3L2, PLA2G2A, TNRSF11B, FGFBP2, MGP, SLPI, DCN, MT-ND2, MTCYB, ADIRF, FRZB, CLEC3A, UPP1, S100A2, PRG4, COL2A1, SOD2 and MT2A. Protein and mRNA expression of MGST1, vimentin, SOD2 and SYF2 (p29) genes validated our scRNA-seq findings. Our data provide new insights into disc cells phenotypes and biomarkers of IVD degeneration that could improve diagnostic and therapeutic options.

Toll-like receptor involvement in adolescent scoliotic facet joint degeneration.

Facet joint osteoarthritis is prevalent in young patients with adolescent idiopathic scoliosis (AIS) and might contribute to back pain. Toll-like receptors (TLR) have been linked to cartilaginous tissue degeneration but their involvement in facet joint osteoarthritis in AIS patients is still unknown. We compared baseline gene expression levels of TLRs -1, -2, -4, and -6 in scoliotic and non-scoliotic chondrocytes and found higher expression levels in scoliotic chondrocytes with significantly higher TLR2 levels. Furthermore, TLR expression correlated strongly and significantly with inflammatory and catabolic markers in scoliotic but not in non-scoliotic chondrocytes. TLR activation with a synthetic TLR2/6 agonist resulted in a robust induction and release of pro-inflammatory and catabolic factors which exacerbated proteoglycan loss in scoliotic but not in non-scoliotic cartilage. We also detected a higher abundance of alarmins including S100A8/9 and biglycan in scoliotic cartilage. Finally, the small-molecule antagonists Sparstolonin B and o-Vanillin reduced catabolism following induction with naturally occurring alarmins and the synthetic TLR2/6 agonist. The high baseline expression, robust responsiveness and strong and significant correlation with proteases and pro-inflammatory cytokines suggest that TLRs are key regulators of facet joint degeneration in AIS. Blocking their activity could therefore potentially modify disease progression.

Low-dose zoledronate for the treatment of bone metastasis secondary to prostate cancer.

BACKGROUND: Bisphosphonates (BPs) including zoledronate (zol) have become standard care for bone metastases as they effectively inhibit tumor-induced osteolysis and associated pain. Several studies have also suggested that zol has direct anti-tumor activity. Systemic administration at high doses is the current approach to deliver zol, yet it has been associated with debilitating side effects. Local therapeutic delivery offers the ability to administer much lower total dosage, while at the same time maintaining sustained high-local drug concentration directly at the target treatment site. Here, we aimed to assess effects of lower doses of zol on bone metastases over a longer time. METHODS: Prostate cancer cell line LAPC4 and prostate-induced bone metastasis cells were treated with zol at 1, 3 and 10 µM for 7 days. Following treatment, cell proliferation was assessed using Almarblue®, Vybrant MTT®, and Live/Dead® viability/cytotoxicity assays. Additionally, cell migration and invasion were carried out using Falcon™ cell culture inserts and Cultrex® 3D spheroid cell invasion assays respectively. RESULTS: We show that treatment with 3-10 µM zol over 7-days significantly decreased cell proliferation in both the prostate cancer cell line LAPC4 and cells from spine metastases secondary to prostate cancer. Using the same low-dose and longer time course for treatment, we demonstrate that 10 µM zol also significantly inhibits tumor cell migration and 3D-cell growth/invasion. CONCLUSIONS: This project harnesses the potential of using zol at low doses for longer treatment periods, which may be a viable treatment modality when coupled with biomaterials or biodevices for local delivery.

Low back pain and disc degeneration are decreased following chronic toll-like receptor 4 inhibition in a mouse model.

OBJECTIVE: Intervertebral disc degeneration is a leading cause of chronic low back pain (LBP) but current treatment is limited. Toll-like receptors (TLRs) on disc cells are activated by endogenous extracellular matrix (ECM) fragments and modulate degeneration in vitro. This study investigated whether inhibiting TLR4 slows disc degeneration and reduces behavioral signs of LBP in vivo. DESIGN: 7-9-month old wild-type and secreted protein acidic and rich in cysteine (SPARC)-null (a model of disc degeneration and LBP) male mice were treated with TAK-242 (TLR4 inhibitor) once, and following a 10-day washout, mice were treated 3 times/week for 8 weeks. Behavioral signs of axial discomfort and radiating leg pain were assessed weekly with the grip force assay and acetone test, respectively. Following treatment, pain-related spinal cord changes were evaluated and lumbar discs were excised and cultured. Cytokine secretion from discs was evaluated with protein arrays. RESULTS: SPARC-null mice displayed elevated signs of axial and radiating pain at baseline compared to wild-type. Chronic, but not acute, TLR4 inhibition reduced behavioral signs of pain compared to vehicle. SPARC-null mice have increased calcitonin gene-related peptide (CGRP)- and glial fibrillary acidic protein (GFAP)-immunoreactivity (astrocyte marker) in the dorsal horn compared to wild-type, which is reduced by chronic TLR4 inhibition. Ex vivo degenerating discs from SPARC-null mice secrete increased levels of many pro-inflammatory cytokines, which chronic TLR4 inhibition reduced. CONCLUSION: Chronic TLR4 inhibition decreased behavioral signs of LBP, pain-related neuroplasticity and disc inflammation in SPARC-null mice. TAK-242 inhibits TLR4 activation within discs, as evidenced by decreases in cytokine release. Therefore, TLRs are potential therapeutic targets to slow disc degeneration and reduce pain.

Nerve Growth Factor Is Regulated by Toll-Like Receptor 2 in Human Intervertebral Discs.

Nerve growth factor (NGF) contributes to the development of chronic pain associated with degenerative connective tissue pathologies, such as intervertebral disc degeneration and osteoarthritis. However, surprisingly little is known about the regulation of NGF in these conditions. Toll-like receptors (TLR) are pattern recognition receptors classically associated with innate immunity but more recently were found to be activated by endogenous alarmins such as fragmented extracellular matrix proteins found in degenerating discs or cartilage. In this study we investigated if TLR activation regulates NGF and which signaling mechanisms control this response in intervertebral discs. TLR2 agonists, TLR4 agonists, or IL-1ß (control) treatment increased NGF, brain-derived neurotrophic factor (BDNF), and IL-1ß gene expression in human disc cells isolated from healthy, pain-free organ donors. However, only TLR2 activation or IL-1ß treatment increased NGF protein secretion. TLR2 activation increased p38, ERK1/2, and p65 activity and increased p65 translocation to the cell nucleus. JNK activity was not affected by TLR2 activation. Inhibition of NF-κB, and to a lesser extent p38, but not ERK1/2 activity, blocked TLR2-driven NGF up-regulation at both the transcript and protein levels. These results provide a novel mechanism of NGF regulation in the intervertebral disc and potentially other pathogenic connective tissues. TLR2 and NF-κB signaling are known to increase cytokines and proteases, which accelerate matrix degradation. Therefore, TLR2 or NF-κB inhibition may both attenuate chronic pain and slow the degenerative progress in vivo.

Photocleavable Hydrogel-Coated Upconverting Nanoparticles: A Multifunctional Theranostic Platform for NIR Imaging and On-Demand Macromolecular Delivery.

Lanthanide-doped upconverting nanoparticles (UCNPs) have emerged as excellent nanotransducers for converting longer wavelength near-infrared (NIR) light to shorter wavelengths spanning the ultraviolet (UV) to the visible (Vis) regions of the spectrum via a multiphoton absorption process, known as upconversion. Here, we report the development of NIR to UV-Vis-NIR UCNPs consisting of LiYF4:Yb(3+)/Tm(3+)@SiO2 individually coated with a 10 ± 2 nm layer of chitosan (CH) hydrogel cross-linked with a photocleavable cross-linker (PhL). We encapsulated fluorescent-bovine serum albumin (FITC-BSA) inside the gel. Under 980 nm excitation, the upconverted UV emission cleaves the PhL cross-links and instantaneously liberates the FITC-BSA under 2 cm thick tissue. The release is immediately arrested if the excitation source is switched off. The upconverted NIR light allows for the tracking of particles under the tissue. Nucleus pulposus (NP) cells cultured with UCNPs are viable both in the presence and in the absence of laser irradiation. Controlled drug delivery of large biomolecules and deep tissue imaging make this system an excellent theranostic platform for tissue engineering, biomapping, and cellular imaging applications.

Axial T1ρ MRI as a diagnostic imaging modality to quantify proteoglycan concentration in degenerative disc disease.

PURPOSE: The aim of the study was to investigate if axial T1ρ MR images had similar accuracy as established sagittal T1ρ MRI for the assessment of proteoglycan concentration and content in intervertebral degenerated discs (IDDs). METHODS: T1ρ and T2-weighted MR images of 12 intervertebral discs (IVDs) from 3 harvested human lumbar spines (levels L1-L2 to L5-S1) were grouped across their degenerative grade (Pfirrmann scores) and analyzed using a 3T MRI scanner in the axial and sagittal views. Post-processing of axial T1ρ-weighted images was performed using a Wiener filter. Median axial T1ρ values for traced regions of interest (ROIs) on color maps were compared against ROIs in the corresponding location in the sagittal plane of each disc. Assessment of sulfated glycosaminoglycans (GAGs) content was also performed. RESULTS: Comparison of post Wiener filtered mid-axial T1ρ values in the NP with corresponding mid-sagittal values revealed no statistical difference (P > 0.05). Higher axial T1ρ and biochemically measured GAGs content corresponded to a lower Pfirrmann grading of the IVDs. A strong association between the T1ρ values and the GAG contents was observed (r = 0.85, P = 0.0002). CONCLUSIONS: The axial T1ρ methodology was validated against sagittal T1ρ providing an augmented spatial representation of IVD and can facilitate localization of focal degeneration within IVDs. T1ρ values provided a better granularity assessment of degenerative disc disease as it correlated with proteoglycan concentration. Thus, Wiener filtering is an effective tool for removing noise from T1ρ-weighted axial MR images.

3D-Printed ABS and PLA Scaffolds for Cartilage and Nucleus Pulposus Tissue Regeneration.

Painful degeneration of soft tissues accounts for high socioeconomic costs. Tissue engineering aims to provide biomimetics recapitulating native tissues. Biocompatible thermoplastics for 3D printing can generate high-resolution structures resembling tissue extracellular matrix. Large-pore 3D-printed acrylonitrile butadiene styrene (ABS) and polylactic acid (PLA) scaffolds were compared for cell ingrowth, viability, and tissue generation. Primary articular chondrocytes and nucleus pulposus (NP) cells were cultured on ABS and PLA scaffolds for three weeks. Both cell types proliferated well, showed high viability, and produced ample amounts of proteoglycan and collagen type II on both scaffolds. NP generated more matrix than chondrocytes; however, no difference was observed between scaffold types. Mechanical testing revealed sustained scaffold stability. This study demonstrates that chondrocytes and NP cells can proliferate on both ABS and PLA scaffolds printed with a simplistic, inexpensive desktop 3D printer. Moreover, NP cells produced more proteoglycan than chondrocytes, irrespective of thermoplastic type, indicating that cells maintain individual phenotype over the three-week culture period. Future scaffold designs covering larger pore sizes and better mimicking native tissue structure combined with more flexible or resorbable materials may provide implantable constructs with the proper structure, function, and cellularity necessary for potential cartilage and disc tissue repair in vivo.

Painful, degenerating intervertebral discs up-regulate neurite sprouting and CGRP through nociceptive factors.

Intervertebral disc degeneration (IVD) can result in chronic low back pain, a common cause of morbidity and disability. Inflammation has been associated with IVD degeneration, however the relationship between inflammatory factors and chronic low back pain remains unclear. Furthermore, increased levels of nerve growth factor (NGF) and brain derived neurotrophic factor (BDNF) are both associated with inflammation and chronic low back pain, but whether degenerating discs release sufficient concentrations of factors that induce nociceptor plasticity remains unclear. Degenerating IVDs from low back pain patients and healthy, painless IVDs from human organ donors were cultured ex vivo. Inflammatory and nociceptive factors released by IVDs into culture media were quantified by enzyme-linked immunosorbent assays and protein arrays. The ability of factors released to induce neurite growth and nociceptive neuropeptide production was investigated. Degenerating discs release increased levels of tumour necrosis factor-α, interleukin-1ß, NGF and BDNF. Factors released by degenerating IVDs increased neurite growth and calcitonin gene-related peptide expression, both of which were blocked by anti-NGF treatment. Furthermore, protein arrays found increased levels of 20 inflammatory factors, many of which have nociceptive effects. Our results demonstrate that degenerating and painful human IVDs release increased levels of NGF, inflammatory and nociceptive factors ex vivo that induce neuronal plasticity and may actively diffuse to induce neo-innervation and pain in vivo.

The C-terminal peptide of chondroadherin modulates cellular activity by selectively binding to heparan sulfate chains.

Chondroadherin, a leucine-rich repeat family member, contains a very C-terminal sequence CKFPTKRSKKAGRH(359), now shown to bind to heparin with a K(D) of 13 µm. This observation led us to investigate whether chondroadherin interacts via this C-terminal heparin-binding domain with glycosaminoglycan chains of proteoglycans at the cell surface. Cells were shown to bind this heparin-binding peptide in FACS analysis, and the interaction was shown to be with glycosaminoglycans because it was abolished when sulfation was inhibited by chlorate treatment of the cells. In separate experiments, heparin and heparan sulfate inhibited the peptide interaction in a dose-dependent manner. Using a human chondrosarcoma and a murine osteoblast cell line, heparan sulfate proteoglycans were identified as the cell surface receptors involved in the binding. Different binding syndecans were identified in the two different cell lines, indicating that the same protein core of a proteoglycan may have structural and functional differences in the attached heparan sulfate chains. Upon binding to coated peptide, cells spread, demonstrating engagement of the cytoskeleton, but no focal adhesion complex was formed. The number of cells adhering via their ß(1) integrin receptor to collagen type II or chondroadherin was profoundly and rapidly enhanced by the addition of the heparin-binding peptide. The peptide added to the cells caused ERK phosphorylation, showing that it triggered intracellular signaling. The results show that heparan sulfate chains differ between various members of the proteoglycan families on a given cell, but also differ between the same proteoglycan on different cells with a potential for differential regulation of cellular activities.

Chondroadherin fragmentation mediated by the protease HTRA1 distinguishes human intervertebral disc degeneration from normal aging.

Chondroadherin, a member of the leucine-rich repeat family, has previously been demonstrated to be fragmented in some juveniles with idiopathic scoliosis. This observation led us to investigate adults with disc degeneration. Immunoblotting analysis demonstrated that non-degenerate discs from three different age groups show no chondroadherin fragmentation. Furthermore, the chondroadherin fragments in adult degenerate disc and the juvenile scoliotic disc were compared via immunoblot analysis and appeared to have a similar size. We then investigated whether or not chondroadherin fragmentation increases with the severity of disc degeneration. Three different samples with different severities were chosen from the same disc, and chondroadherin fragmentation was found to be more abundant with increasing severity of degeneration. This observation led us to the creation of a neoepitope antibody to the cleavage site observed. We then observed that the cleavage site in adult degenerate discs and juvenile scoliotic discs was identical as confirmed by the neoepitope antibody. Consequently, investigation of the protease capable of cleaving chondroadherin at this site was necessary. In vitro digests of disc tissue demonstrated that ADAMTS-4 and -5; cathepsins K, B, and L; and MMP-3, -7, -12, and -13 were incapable of cleavage of chondroadherin at this site and that HTRA1 was indeed the only protease capable. Furthermore, increased protein levels of the processed form of HTRA1 were demonstrated in degenerate disc tissues via immunoblotting. The results suggest that chondroadherin fragmentation can be used as a biomarker to distinguish the processes of disc degeneration from normal aging.

Low-frequency high-magnitude mechanical strain of articular chondrocytes activates p38 MAPK and induces phenotypic changes associated with osteoarthritis and pain.

Osteoarthritis (OA) is a debilitating joint disorder resulting from an incompletely understood combination of mechanical, biological, and biochemical processes. OA is often accompanied by inflammation and pain, whereby cytokines associated with chronic OA can up-regulate expression of neurotrophic factors such as nerve growth factor (NGF). Several studies suggest a role for cytokines and NGF in OA pain, however the effects of changing mechanical properties in OA tissue on chondrocyte metabolism remain unclear. Here, we used high-extension silicone rubber membranes to examine if high mechanical strain (HMS) of primary articular chondrocytes increases inflammatory gene expression and promotes neurotrophic factor release. HMS cultured chondrocytes displayed up-regulated NGF, TNFα and ADAMTS4 gene expression while decreasing TLR2 expression, as compared to static controls. HMS culture increased p38 MAPK activity compared to static controls. Conditioned medium from HMS dynamic cultures, but not static cultures, induced significant neurite sprouting in PC12 cells. The increased neurite sprouting was accompanied by consistent increases in PC12 cell death. Low-frequency high-magnitude mechanical strain of primary articular chondrocytes in vitro drives factor secretion associated with degenerative joint disease and joint pain. This study provides evidence for a direct link between cellular strain, secretory factors, neo-innervation, and pain in OA pathology.

Exploring the Impact of TLR-2 Signaling on miRNA Dysregulation in Intervertebral Disc Degeneration.

Toll-like receptors (TLRs) are key mediators of inflammation in intervertebral disc (IVD) degeneration. TLR-2 activation contributes to the degenerative process by increasing the expression of extracellular matrix-degrading enzymes, pro-inflammatory cytokines, and neurotrophins. As potent post-transcriptional regulators, microRNAs can modulate intracellular mechanisms, and their dysregulation is known to contribute to numerous pathologies. This study aims to investigate the impact of TLR-2 signaling on miRNA dysregulation in the context of IVD degeneration. Small-RNA sequencing of degenerated IVD cells shows the dysregulation of ten miRNAs following TLR-2 activation by PAM2CSK4. The miR-155-5p is most significantly upregulated in degenerated and non-degenerated annulus fibrosus and nucleus pulposus cells. Sequence-based target and pathway prediction shows the involvement of miR-155-5p in inflammation- and cell fate-related pathways and TLR-2-induced miR-155-5p expression leads to the downregulation of its target c-FOS. Furthermore, changes specific to the activation of TLR-2 through fragmented fibronectin are seen in miR-484 and miR-487. Lastly, miR-100-3p, miR-320b, and miR-181a-3p expression exhibit degeneration-dependent changes. These results show that TLR-2 signaling leads to the dysregulation of miRNAs in IVD cells as well as their possible downstream effects on inflammation and degeneration. The identified miRNAs provide important opportunities as potential therapeutic targets for IVD degeneration and low back pain.

Injectable hydrogel induces regeneration of naturally degenerate human intervertebral discs in a loaded organ culture model.

Low back pain resulting from disc degeneration is a leading cause of disability worldwide. However, to date few therapies target the cause and fail to repair the intervertebral disc (IVD). This study investigates the ability of an injectable hydrogel (NPgel), to inhibit catabolic protein expression and promote matrix expression in human nucleus pulposus (NP) cells within a tissue explant culture model isolated from degenerate discs. Furthermore, the injection capacity of NPgel into naturally degenerate whole human discs, effects on mechanical function, and resistance to extrusion during loading were investigated. Finally, the induction of potential regenerative effects in a physiologically loaded human organ culture system was investigated following injection of NPgel with or without bone marrow progenitor cells. Injection of NPgel into naturally degenerate human IVDs increased disc height and Young's modulus, and was retained during extrusion testing. Injection into cadaveric discs followed by culture under physiological loading increased MRI signal intensity, restored natural biomechanical properties and showed evidence of increased anabolism and decreased catabolism with tissue integration observed. These results provide essential proof of concept data supporting the use of NPgel as an injectable therapy for disc regeneration. STATEMENT OF SIGNIFICANCE: Low back pain resulting from disc degeneration is a leading cause of disability worldwide. However, to date few therapies target the cause and fail to repair the intervertebral disc. This study investigated the potential regenerative properties of an injectable hydrogel system (NPgel) within human tissue samples. To mimic the human in vivo conditions and the unique IVD niche, a dynamically loaded intact human disc culture system was utilised. NPgel improved the biomechanical properties, increased MRI intensity and decreased degree of degeneration. Furthermore, NPgel induced matrix production and decreased catabolic factors by the native cells of the disc. This manuscript provides evidence for the potential use of NPgel as a regenerative biomaterial for intervertebral disc degeneration.

Markers of Tissue Deterioration and Pain on Earth and in Space.

Purpose: Pain is an understudied physiological effect of spaceflight. Changes in inflammatory and tissue degradation markers are often associated with painful conditions. Our aim was to evaluate the changes in markers associated with tissue deterioration after a short-term spaceflight. Patients and Methods: Plasma levels of markers for systemic inflammation and tissue degeneration markers were assessed in two astronauts before and within 24 h after the 17-day Axiom Space AX-1 mission. Results: After the spaceflight, C-reactive protein (CRP) was reduced in both astronauts, while INFγ, GM-CSF, TNFα, BDNF, and all measured interleukins were consistently increased. Chemokines demonstrated variable changes, with consistent positive changes in CCL3, 4, 8, 22 and CXCL8, 9, 10, and consistent negative change in CCL8. Markers associated with tissue degradation and bone turnover demonstrated consistent increases in MMP1, MMP13, NTX and OPG, and consistent decreases in MMP3 and MMP9. Conclusion: Spaceflight induced changes in the markers of systemic inflammation, tissue deterioration, and bone resorption in two astronauts after a short, 17-day, which were often consistent with those observed in painful conditions on Earth. However, some differences, such as a consistent decrease in CRP, were noted. All records for the effect of space travel on human health are critical for improving our understanding of the effect of this unique environment on humans.

Viscoelastic hydrogels regulate adipose-derived mesenchymal stem cells for nucleus pulposus regeneration.

Low back pain is a leading cause of disability worldwide, often attributed to intervertebral disc (IVD) degeneration with loss of the functional nucleus pulposus (NP). Regenerative strategies utilizing biomaterials and stem cells are promising for NP repair. Human NP tissue is highly viscoelastic, relaxing stress rapidly under deformation. However, the impact of tissue-specific viscoelasticity on the activities of adipose-derived stem cells (ASC) remains largely unexplored. Here, we investigated the role of matrix viscoelasticity in regulating ASC differentiation for IVD regeneration. Viscoelastic alginate hydrogels with stress relaxation time scales ranging from 100 s to 1000s were developed and used to culture human ASCs for 21 days. Our results demonstrated that the fast-relaxing hydrogel significantly enhanced ASCs long-term cell survival and NP-like extracellular matrix secretion of aggrecan and type-II collagen. Moreover, gene expression analysis revealed a substantial upregulation of the mechanosensitive ion channel marker TRPV4 and NP-specific markers such as SOX9, HIF-1α, KRT18, CDH2 and CD24 in ASCs cultured within the fast-relaxing hydrogel, compared to slower-relaxing hydrogels. These findings highlight the critical role of matrix viscoelasticity in regulating ASC behavior and suggest that viscoelasticity is a key parameter for novel biomaterials design to improve the efficacy of stem cell therapy for IVD regeneration. STATEMENT OF SIGNIFICANCE: Systematically characterized the influence of tissue-mimetic viscoelasticity on ASC. NP-mimetic hydrogels with tunable viscoelasticity and tissue-matched stiffness. Long-term survival and metabolic activity of ASCs are substantially improved in the fast-relaxing hydrogel. The fast-relaxing hydrogel allows higher rate of cell protrusions formation and matrix remodeling. ASC differentiation towards an NP-like cell phenotype is promoted in the fast-relaxing hydrogel, with more CD24 positive expression indicating NP committed cell fate. The expression of TRPV4, a molecular sensor of matrix viscoelasticity, is significantly enhanced in the fast-relaxing hydrogel, indicating ASC sensing matrix viscoelasticity during cell development. The NP-specific ECM secretion of ASC is considerably influenced by matrix viscoelasticity, where the deposition of aggrecan and type-II collagen are significantly enhanced in the fast-relaxing hydrogel.

Impact of the COVID-19 pandemic on the productivity and career prospects of musculoskeletal researchers.

Academic researchers faced a multitude of challenges posed by the COVID-19 pandemic, including widespread shelter-in-place orders, workplace closures, and cessation of in-person meetings and laboratory activities. The extent to which these challenges impacted musculoskeletal researchers, specifically, is unknown. We developed an anonymous web-based survey to determine the pandemic's impact on research productivity and career prospects among musculoskeletal research trainees and faculty. There were 116 musculoskeletal (MSK) researchers with varying demographic backgrounds who completed the survey. Of respondents, 48.3% (n = 56) believed that musculoskeletal funding opportunities decreased because of COVID-19, with faculty members more likely to hold this belief compared to nonfaculty researchers (p = 0.008). Amongst MSK researchers, 88.8% (n = 103) reported research activity was limited by COVID-19, and 92.2% (n = 107) of researchers reported their research was not able to be refocused on COVID-19-related topics, with basic science researchers less likely to be able to refocus their research compared to clinical researchers (p = 0.030). Additionally, 47.4% (n = 55) reported a decrease in manuscript submissions since the onset of the pandemic. Amongst 51 trainee researchers, 62.8% (n = 32) reported a decrease in job satisfaction directly attributable to the COVID-19 pandemic. In summary, study findings indicated that MSK researchers struggled to overcome challenges imposed by the pandemic, reporting declines in funding opportunities, research productivity, and manuscript submission. Trainee researchers experienced significant disruptions to critical research activities and worsening job satisfaction. Our findings motivate future efforts to support trainees in developing their careers and target the recovery of MSK research from the pandemic stall.