Non-viral precision T cell receptor replacement for personalized cell therapy.

T cell receptors (TCRs) enable T cells to specifically recognize mutations in cancer cells1-3. Here we developed a clinical-grade approach based on CRISPR-Cas9 non-viral precision genome-editing to simultaneously knockout the two endogenous TCR genes TRAC (which encodes TCRα) and TRBC (which encodes TCRß). We also inserted into the TRAC locus two chains of a neoantigen-specific TCR (neoTCR) isolated from circulating T cells of patients. The neoTCRs were isolated using a personalized library of soluble predicted neoantigen-HLA capture reagents. Sixteen patients with different refractory solid cancers received up to three distinct neoTCR transgenic cell products. Each product expressed a patient-specific neoTCR and was administered in a cell-dose-escalation, first-in-human phase I clinical trial ( NCT03970382 ). One patient had grade 1 cytokine release syndrome and one patient had grade 3 encephalitis. All participants had the expected side effects from the lymphodepleting chemotherapy. Five patients had stable disease and the other eleven had disease progression as the best response on the therapy. neoTCR transgenic T cells were detected in tumour biopsy samples after infusion at frequencies higher than the native TCRs before infusion. This study demonstrates the feasibility of isolating and cloning multiple TCRs that recognize mutational neoantigens. Moreover, simultaneous knockout of the endogenous TCR and knock-in of neoTCRs using single-step, non-viral precision genome-editing are achieved. The manufacture of neoTCR engineered T cells at clinical grade, the safety of infusing up to three gene-edited neoTCR T cell products and the ability of the transgenic T cells to traffic to the tumours of patients are also demonstrated.

Neoantigen-targeted CD8+ T cell responses with PD-1 blockade therapy.

Neoantigens are peptides derived from non-synonymous mutations presented by human leukocyte antigens (HLAs), which are recognized by antitumour T cells1-14. The large HLA allele diversity and limiting clinical samples have restricted the study of the landscape of neoantigen-targeted T cell responses in patients over their treatment course. Here we applied recently developed technologies15-17 to capture neoantigen-specific T cells from blood and tumours from patients with metastatic melanoma with or without response to anti-programmed death receptor 1 (PD-1) immunotherapy. We generated personalized libraries of neoantigen-HLA capture reagents to single-cell isolate the T cells and clone their T cell receptors (neoTCRs). Multiple T cells with different neoTCR sequences (T cell clonotypes) recognized a limited number of mutations in samples from seven patients with long-lasting clinical responses. These neoTCR clonotypes were recurrently detected over time in the blood and tumour. Samples from four patients with no response to anti-PD-1 also demonstrated neoantigen-specific T cell responses in the blood and tumour to a restricted number of mutations with lower TCR polyclonality and were not recurrently detected in sequential samples. Reconstitution of the neoTCRs in donor T cells using non-viral CRISPR-Cas9 gene editing demonstrated specific recognition and cytotoxicity to patient-matched melanoma cell lines. Thus, effective anti-PD-1 immunotherapy is associated with the presence of polyclonal CD8+ T cells in the tumour and blood specific for a limited number of immunodominant mutations, which are recurrently recognized over time.

Complete resorption of the humerus in metastatic thyroid carcinoma: a case report.

BACKGROUND: Thyroid carcinoma is the most common endocrinological malignancy, but its spread to bone is rare. Particularly, bone metastases leading to complete resorption of the humerus are extremely uncommon. We aimed to explore factors affecting treatment decision in humeral metastasis by presenting a case and analyze the possible treatments via conducting a literature review. CASE PRESENTATION: We described a case of a 68-year-old woman experiencing chronic pain in her right upper arm for six years. Clinical, radiological, and pathological evaluations confirmed humeral metastasis from thyroid carcinoma. Surgical treatments like tumor removal or limb amputation were suggested for prolonging life and pain relief, but the patient refused them and pursued conservative managements such as herbal medicine, radioactive iodine (131I) therapy, and Levothyroxine Sodium(L-T4). The humeral destruction aggravated gradually, ultimately leading to complete resorption of her right humerus. The patient could not move her right shoulder, but her forearm motion was almost normal; thus, she could complete most of her daily living activities independently. Surgical treatments such as limb amputation were advised but she still refused them for preservation of the residual limb function and preferred conservative managements. CONCLUSION: A personalized multidisciplinary approach is important for patients with bone metastasis. The balance between limb amputation for life-prolonging and pain relief and limb salvage for preservation of residual function and social and psychological well-being should be considered. Our literature review revealed that some novel surgical treatments and techniques are available for bone metastases. This case adds to our current understanding of bone metastases and will contribute to future research and treatments.

Engineering stem cells to produce exosomes with enhanced bone regeneration effects: an alternative strategy for gene therapy.

BACKGROUND: Exosomes derived from stem cells have been widely studied for promoting regeneration and reconstruction of multiple tissues as "cell-free" therapies. However, the applications of exosomes have been hindered by limited sources and insufficient therapeutic potency. RESULTS: In this study, a stem cell-mediated gene therapy strategy is developed in which mediator mesenchymal stem cells are genetically engineered by bone morphogenetic protein-2 gene to produce exosomes (MSC-BMP2-Exo) with enhanced bone regeneration potency. This effect is attributed to the synergistic effect of the content derived from MSCs and the up-regulated BMP2 gene expression. The MSC-BMP2-Exo also present homing ability to the injured site. The toxic effect of genetical transfection vehicles is borne by mediator MSCs, while the produced exosomes exhibit excellent biocompatibility. In addition, by plasmid tracking, it is interesting to find a portion of plasmid DNA can be encapsulated by exosomes and delivered to recipient cells. CONCLUSIONS: In this strategy, engineered MSCs function as cellular factories, which effectively produce exosomes with designed and enhanced therapeutic effects. The accelerating effect in bone healing and the good biocompatibility suggest the potential clinical application of this strategy.

A novel surgical planning system using an AI model to optimize planning of pedicle screw trajectories with highest bone mineral density and strongest pull-out force.

OBJECTIVE: The purpose of this study was to evaluate the ability of a novel artificial intelligence (AI) model in identifying optimized transpedicular screw trajectories with higher bone mineral density (BMD) as well as higher pull-out force (POF) in osteoporotic patients. METHODS: An innovative pedicle screw trajectory planning system called Bone's Trajectory was developed using a 3D graphic search and an AI-based finite element analysis model. The preoperative CT scans of 21 elderly osteoporotic patients were analyzed retrospectively. The AI model automatically calculated the number of alternative transpedicular trajectories, the trajectory BMD, and the estimated POF of L3-5. The highest BMD and highest POF of optimized trajectories were recorded and compared with AO standard trajectories. RESULTS: The average patient age and average BMD of the vertebral bodies were 69.6 ± 7.8 years and 55.9 ± 17.1 mg/ml, respectively. On both sides of L3-5, the optimized trajectories showed significantly higher BMD and POF than the AO standard trajectories (p < 0.05). On average, the POF of optimized trajectory screws showed at least a 2.0-fold increase compared with AO trajectory screws. CONCLUSIONS: The novel AI model performs well in enabling the selection of optimized transpedicular trajectories with higher BMD and POF than the AO standard trajectories.

Osteoarthritis Pain.

Joint pain is the hallmark symptom of osteoarthritis (OA) and the main reason for patients to seek medical assistance. OA pain greatly contributes to functional limitations of joints and reduced quality of life. Although several pain-relieving medications are available for OA treatment, the current intervention strategy for OA pain cannot provide satisfactory pain relief, and the chronic use of the drugs for pain management is often associated with significant side effects and toxicities. These observations suggest that the mechanisms of OA-related pain remain undefined. The current review mainly focuses on the characteristics and mechanisms of OA pain. We evaluate pathways associated with OA pain, such as nerve growth factor (NGF)/tropomyosin receptor kinase A (TrkA), calcitonin gene-related peptide (CGRP), C-C motif chemokine ligands 2 (CCL2)/chemokine receptor 2 (CCR2) and tumor necrosis factor alpha (TNF-α), interleukin-1beta (IL-1ß), the NOD-like receptor (NLR) family, pyrin domain-containing protein 3 (NLRP3) inflammasome, and the Wnt/ß-catenin signaling pathway. In addition, animal models currently used for OA pain studies and emerging preclinical studies are discussed. Understanding the multifactorial components contributing to OA pain could provide novel insights into the development of more specific and effective drugs for OA pain management.

Sleep Deprivation Affects Tau Phosphorylation in Human Cerebrospinal Fluid.

Tau hyperphosphorylation is an early step in tau-mediated neurodegeneration and is associated with intracellular aggregation of tau as neurofibrillary tangles, neuronal and synaptic loss, and eventual cognitive dysfunction in Alzheimer disease. Sleep loss increases the cerebrospinal fluid concentration of amyloid-ß and tau. Using mass spectrometry, we measured tau and phosphorylated tau concentrations in serial samples of cerebrospinal fluid collected from participants who were sleep-deprived, treated with sodium oxybate, or allowed to sleep normally. We found that sleep loss affected phosphorylated tau differently depending on the modified site. These findings suggest a mechanism for sleep loss to increase risk of Alzheimer disease. ANN NEUROL 2020;87:700-709.

Metformin Hydrochloride Encapsulation by Alginate Strontium Hydrogel for Cartilage Regeneration by Reliving Cellular Senescence.

Cartilage lesion is a common tissue defect and is challenging in clinical practice. Trauma-induced cellular senescence could decrease the chondrocyte capability of maintaining cartilage tissue regeneration. A previous investigation showed that, by controlling the cellular senescence, the cartilage regeneration can be significantly accelerated. Based on this finding, we design a novel hydrogel, Alg/MH-Sr, that combines metformin, an established drug for inhibiting senescence, and strontium, an effective anti-inflammatory material for cartilage tissue engineering. A RT-PCR test suggests the significant inhibitory effect of the hydrogel on senescent, apoptotic, oxidative, and inflammatory genes' expression. Histological examinations demonstrate that the Alg/MH-Sr hydrogel accelerated cartilage repairment, and chondrocyte senescence was significantly inhibited. Our study demonstrates that the Alg/MH-Sr hydrogel is effective for cartilage defect treatment and provides a new clue in accelerating tissue repairment by inhibiting the senescence of cells and tissues.

Proximal Tibial Cortex Transverse Distraction Facilitating Healing and Limb Salvage in Severe and Recalcitrant Diabetic Foot Ulcers.

BACKGROUND: The management of severe and recalcitrant diabetic foot ulcers is challenging. Distraction osteogenesis is accompanied by vascularization and regeneration of the surrounding tissues. Longitudinal distraction of the proximal tibia stimulates increased and prolonged blood flow to the distal tibia. However, the effects of transverse distraction of the proximal tibia cortex on severe and recalcitrant diabetic foot ulcers are largely unknown. QUESTIONS/PURPOSES: (1) Does tibial cortex transverse distraction increase healing and decrease major amputation and recurrence of severe and recalcitrant diabetic foot ulcers compared with routine management (which generally included débridement, revascularization, negative pressure wound therapy, local or free flaps, or skin grafts as indicated)? (2) Does neovascularization and perfusion increase at the foot after the procedure? (3) What are the complications of tibial cortex transverse distraction in patients with severe and recalcitrant diabetic foot ulcers? METHODS: Between July 2014 and March 2017, we treated 136 patients with diabetes mellitus and University of Texas Grade 2B to 3D ulcers (wound penetrating to the tendon, capsule, bone, or joint with infection and/or ischemia). The patients had failed to respond to treatment for at least 6 months, and their ulcers had a mean ± SD area of 44 cm ± 10 cm. All 136 patients underwent tibial cortex transverse distraction (partial corticotomy of the upper tibia, which was in normal condition, followed by 4 weeks of transverse distraction medially then laterally). We compared these patients with the last 137 consecutive patients we treated with standard surgical treatment, consisting of débridement, revascularization, local or free flap or skin equivalent, or graft reconstruction along with negative-pressure wound therapy between May 2011 and June 2013; there was a 1-year period during which both treatments were in use, and we did not include patients whose procedures were performed during this time in either group. Patients in both groups received standard off-loading and wound care. The patients lost to follow-up by 2 years (0.7% of the treatment group [one of 137] and 1.4% of the control group [two of 139]; p = 0.57) were excluded. The patients in the treatment and control groups had a mean age of 61 years and 60 years, respectively, and they were predominantly men in both groups (70% [95 of 136] versus 64% [88 of 137]; p = 0.32). There were no differences with respect to parameters associated with the likelihood of ulcer healing, such as diabetes and ulcer duration, ulcer grades and area, smoking, and arterial status. We compared the groups with respect to ulcer healing (complete epithelialization without discharge, maintained for at least 2 weeks, which was determined by an assessor not involved with clinical care) in a 2-year follow-up, the proportion of ulcers that healed by 6 months, major amputation, recurrence, and complications in the 2-year follow-up. Foot arterial status and perfusion were assessed in the tibial cortex transverse distraction group using CT angiography and perfusion imaging. RESULTS: The tibial cortex transverse distraction group had a higher proportion of ulcers that healed in the 2-year follow-up than the control group (96% [131 of 136] versus 68% [98 of 137]; odds ratio 10.40 [95% confidence interval 3.96 to 27.43]; p < 0.001). By 6 months, a higher proportion of ulcers healed in the tibia cortex transverse distraction group than the control group (93% [126 of 136] versus 41% [56 of 137]; OR 18.2 [95% CI 8.80 to 37.76]; p < 0.001). Lower proportions of patients in the tibia cortex transverse distraction group underwent major amputation (2.9% [four of 136] versus 23% [31 of 137], OR 0.10 [95% CI 0.04 to 0.30]; p < 0.001) or had recurrences 2.9% (4 of 136) versus 17% (23 of 137), OR 0.20 [95% CI 0.05 to 0.45]; p < 0.001) than the control group in 2-year follow-up. In the feet of the patients in the tibial cortex transverse distraction group, there was a higher density of small vessels (19 ± 2.1/mm versus 9 ± 1.9/mm; mean difference 10/mm; p = 0.010), higher blood flow (24 ± 5 mL/100 g/min versus 8 ± 2.4 mL/100 g/min, mean difference 16 mL/100 g/min; p = 0.004) and blood volume (2.5 ± 0.29 mL/100 g versus 1.3 ± 0.33 mL/100 g, mean difference 1.2 mL/100 g; p = 0.03) 12 weeks postoperatively than preoperatively. Complications included closed fractures at the corticotomy site (in 1.5% of patients; two of 136), which were treated with closed reduction and healed, as well as pin-site infections (in 2.2% of patients; three of 136), which were treated with dressing changes and they resolved without osteomyelitis. CONCLUSIONS: Proximal tibial cortex transverse distraction substantially facilitated healing and limb salvage and decreased the recurrence of severe and recalcitrant diabetic foot ulcers. The surgical techniques were relatively straightforward although the treatment was unorthodox, and the complications were few and minor. These findings suggest that tibial cortex transverse distraction is an effective procedure to treat severe and recalcitrant diabetic foot ulcers compared with standard surgical therapy. Randomized controlled trials are required to confirm these findings. LEVEL OF EVIDENCE: Level II, therapeutic study.

Impact of credentialing and provider privileges on clinical interventions made by advanced practice pharmacists in California.

BACKGROUND: The Advanced Practice Pharmacist (APh) licensure has provided an opportunity for pharmacists to expand their scope of practice in California; however, there have not been any studies in California assessing the interventions made by APhs as credentialed providers of care. OBJECTIVE: To assess the clinical interventions made by APhs as credentialed providers with clinical privileges in an independent community pharmacy. METHODS: This was a retrospective, observational study that assessed clinical interventions made by APhs on patients referred for disease state management between January 2018 and December 2018. Pharmacist interventions were stratified into 3 levels of provider care: full privilege (FP), limited privilege (LP), and no privilege. RESULTS: FP had the highest percentage of accepted recommendations (62.2% ± 20.1%), whereas LP and no privilege had lower percentages of accepted recommendations (41.9% ± 12.0% and 31.6% ± 3.7%, respectively) (P < 0.01). CONCLUSION: APhs as credentialed providers with FP, or even LP, made more successful clinical interventions than those without any privileges.

Multifunctional Nanoengineered Hydrogels Consisting of Black Phosphorus Nanosheets Upregulate Bone Formation.

Tissue-engineered hydrogels have received extensive attention as their mechanical properties, chemical compositions, and biological signals can be dynamically modified for mimicking extracellular matrices (ECM). Herein, the synthesis of novel double network (DN) hydrogels with tunable mechanical properties using combinatorial screening methods is reported. Furthermore, nanoengineered (NE) hydrogels are constructed by addition of ultrathin 2D black phosphorus (BP) nanosheets to the DN hydrogels with multiple functions for mimicking the ECM microenvironment to induce tissue regeneration. Notably, it is found that the BP nanosheets exhibit intrinsic properties for induced CaP crystal particle formation and therefore improve the mineralization ability of NE hydrogels. Finally, in vitro and in vivo data demonstrate that the BP nanosheets, mineralized CaP crystal nanoparticles, and excellent mechanical properties provide a favorable ECM microenvironment to mediate greater osteogenic cell differentiation and bone regeneration. Consequently, the combination of bioactive chemical materials and excellent mechanical stimuli of NE hydrogels inspire novel engineering strategies for bone-tissue regeneration.

Matrix metallopeptidase 9 targeted by hsa-miR-494 promotes silybin-inhibited osteosarcoma.

Osteosarcoma (OS) is the most common malignant tumor that develops in bone. Its mortality is very high. Therefore, study of mechanisms of pathogenesis of the OS is urgently required. Previous studies of microarray showed that the expression levels of matrix metallopeptidase 9 (MMP-9) altered significantly in OS. In addition, overexpression of MMP-9 is recognized as an indicator in cancer. However, the exact roles of MMP-9 in OS are not fully investigated. Thus, we firstly studied the roles of MMP-9 in OS and revealed that silence of MMP-9 inhibited OS cell proliferation as determined by MTT assay and colony formation assay. Secondly, we conducted TUNEL assay and confirmed loss of functions of MMP-9 induced OS cell apoptosis. Next, we used lentivector packaging method to overexpress MMP-9 and found that overexpression of MMP-9 promoted OS cell migration. Fourthly, the results of luciferase assay showed that MMP-9 was targeted by hsa-miR-494, which inhibited OS. Fifthly, we revealed that the levels of hsa-miR-494 were upregulated by the drug silybin which inhibited OS. Finally, we revealed that silybin inhibited OS cell viability by altering the protein levels of ß-catenin and Runt-related transcription factor 2 (RUNX2) as determined by western blot and immunocytochemistry (ICC).

Aging Reduces an ERRalpha-Directed Mitochondrial Glutaminase Expression Suppressing Glutamine Anaplerosis and Osteogenic Differentiation of Mesenchymal Stem Cells.

Aging deteriorates osteogenic capacity of mesenchymal stem/stromal cells (MSCs), contributing to imbalanced bone remodeling and osteoporosis. Glutaminase (Gls) catabolizes glutamine into glutamate at the first step of mitochondrial glutamine (Gln)-dependent anaplerosis which is essential for MSCs upon osteogenic differentiation. Estrogen-related receptor α (ERRα) regulates genes required for mitochondrial function. Here, we found that ERRα and Gls are upregulated by osteogenic induction in human MSCs (hMSCs). In contrast, osteogenic differentiation capacity and glutamine consumption of MSCs, as well as ERRα, Gls and osteogenic marker genes are significantly reduced with age. We demonstrated that ERRα binds to response elements on Gls promoter and affects glutamine anaplerosis through transcriptional induction of Gls. Conversely, mTOR inhibitor rapamycin, ERRα inverse agonist compound 29 or Gls inhibitor BPTES leads to reduced Gln anaplerosis and deteriorated osteogenic differentiation of hMSCs. Importantly, overexpression of ERRα or Gls restored impairment by these inhibitors. Finally, we proved that compensated ERRα or Gls expression indeed potentiated Gln anaplerosis and osteogenic capability of elderly mice MSCs in vitro. Together, we establish that Gls is a novel ERRα target gene and ERRα/Gls signaling pathway plays an important role in osteogenic differentiation of MSCs, providing new sights into novel regenerative therapeutics development. Our findings suggest that restoring age-related mitochondrial Gln-dependent anaplerosis may be beneficial for degenerative bone disorders such as osteoporosis. Stem Cells 2017;35:411-424.

The association of lumbar intervertebral disc calcification on plain radiographs with the UTE Disc Sign on MRI.

PURPOSE: The pathogenesis and the clinical impact of disc calcification are not well known. Utilizing ultra-short time-to-echo (UTE) magnetic resonance imaging, the UTE Disc Sign (UDS) (i.e., hypo/hyper-intense disc band) was developed and found to be more significantly related to pain and disability than the conventional T2-weighted (T2W) MRI. It has been hypothesized that the UDS may represent mineralized deposits in the disc. The following study addressed the relationship between disc calcification on plain radiographs to that of the UDS on MRI. METHODS: A cross-sectional study was performed on 106 Southern Chinese subjects (50% male; mean age 52.3 years). Standing lateral plain radiographs as well as T2W and UTE MRI of L1-S1 (n = 530 discs) were performed of all subjects. Lateral radiographs were used to localize disc calcification of the lumbar spine, T2W MRI was utilized to assess disc degeneration based on a defined grading scheme, and the UTE MRI was implemented to detect the UDS (hyper- or hypo-intense band across a disc). Disc degeneration and UDS scores were summed to represent cumulative scores. Subject demographics and disability profiles (Oswestry Disability Index: ODI) were obtained. RESULTS: Disc calcification on plain radiographs was observed in 33.9% of subjects (55.5% males; mean age 54.3 years), whereas UDS was noted in 40.5% of subjects (51.1% males; mean age 55.0 years). Of these subjects, 66.6% calcification and 74.4% UDS occurred at the three lowest lumbar levels, while multilevel calcification and UDS involved 19.4 and 39.5%, respectively. 72.2% of subjects with plain radiographic disc calcification had corresponding UDS on UTE MRI (p < 0.001). Multilevel disc calcification on plain radiographs was associated with multilevel UDS (71.4%, p < 0.001). Both the number of calcified disc levels on plain radiographs and the number of UDS levels were also significantly and positively correlated with each other (r = 0.58, p < 0.001). Subjects with disc calcification and positive UDS as well as individuals with increased disc degeneration scores on T2 W MRI were significantly older (p < 0.05). The cumulative UDS score on UTE MRI significantly correlated with worse ODI scores (r = 0.31; p = 0.001), whereas cumulative disc calcification scores on plain radiographs did not (r = 0.15; p = 0.19). CONCLUSIONS: This is the first study to compare the UDS on UTE MRI with disc calcification on plain radiographs. Disc calcification was correlated with the UDS on UTE, suggesting that the UDS may represent disc calcification. However, UTE MRI appears to be a more sensitive imaging modality in identifying subtle and unique disc changes that may not be revealed on plain radiographs or conventional MRI. This disconnect may rationalize the significant correlation of UTE with disability in comparison with the conventional imaging, further stressing its potential clinical importance.

Lumbar intervertebral disc allograft transplantation: the revascularisation pattern.

PURPOSE: Fresh frozen intervertebral disc allograft transplantation has been reported to be a viable treatment option for advanced degenerative disc diseases, but rapid degeneration of the postoperative allograft was found. Loss of nutrient supply is believed to be the most likely inducer because the disc allografts have to endure in an ischaemic environment until the nutrient pathway is re-established. The aim of this study was to focus on the revascularisation of the disc allograft after transplantation in goats. METHODS: Twenty male goats were used in this study. Intervertebral disc allograft transplantation was performed at L4/L5. Groups of five goats were killed at 1.5, 6 and 12 m postoperatively, respectively. The transplanted segments were harvested, fixed, sagittally cut and decalcified for H&E staining and immunochemistry to observe the blood vessel formation at the endplates, anterior outer annulus, posterior outer annulus, inner annulus and the nucleus. The blood vessel density and the sectional vessel area were measured. RESULTS: Blood vessels were first found in the marrow space of the bony endplate and the outer annulus at 1.5 month postoperatively. Then, they were able to penetrate to reach the cartilaginous endplate and the inner annulus after 6 months. Interestingly, the endplate area possessed the most abundant blood vessels, with the highest level of vessel density and area at the final follow-up. None of these newly formed vessels invaded the nucleus during the observation period. CONCLUSIONS: Revascularisation of the postoperative disc allograft has been determined, but its pattern was different from that in adult normal discs, suggesting that the typical nutrient diffusion pattern may be affected after transplantation.

Lumbar intervertebral disc allograft transplantation: long-term mobility and impact on the adjacent segments.

PURPOSE: Fresh-frozen intervertebral disc (IVD) allograft transplantation has been successfully performed in the human cervical spine. Whether this non-fusion technology could truly decrease adjacent segment disease is still unknown. This study evaluated the long-term mobility of the IVD-transplanted segment and the impact on the adjacent spinal segments in a goat model. METHODS: Twelve goats were used. IVD allograft transplantation was performed at lumbar L4/L5 in 5 goats; the other 7 goats were used as the untreated control (5) and for the supply of allografts (2). Post-operation lateral radiographs of the lumbar spine in the neutral, full-flexion and full-extension positions were taken at 1, 3, 6, 9 and 12 months. Disc height (DH) of the allograft and the adjacent levels was calculated and range of motion (ROM) was measured using the Cobb's method. The anatomy of the adjacent discs was observed histologically. RESULTS: DH of the transplanted segment was decreased significantly after 3 months but no further reduction was recorded until the final follow-up. No obvious alteration was seen in the ROM of the transplanted segment at different time points with the ROM at 12 months being comparable to that of the untreated control. The DH and ROM in the adjacent segments were well maintained during the whole observation period. At post-operative 12 months, the ROM of the adjacent levels was similar to that of the untreated control and the anatomical morphology was well preserved. CONCLUSIONS: Lumbar IVD allograft transplantation in goats could restore the segmental mobility and did not negatively affect the adjacent segments after 12 months.

Osseointegrated Prosthetic Limb for the treatment of lower limb amputations : Experience and outcomes.

BACKGROUND: Osseointegration has emerged over the past two decades as a dramatically different approach for the treatment of lower limb amputations, which involves direct attachment of the prosthesis to the skeletal residuum. This approach can address many of the socket-interface issues associated with socket prostheses which represent the current standard of care for amputees. The Osseointegrated Prosthetic Limb (OPL) is an osseointegration implant with a new design and improved features compared to other available implant systems. OBJECTIVES: To report on the experience and outcomes of using the OPL for osseointegrated reconstruction of lower limb amputations. MATERIALS AND METHODS: This is a retrospective study of 22 patients who received the OPL implant between December 2013 and November 2014. Clinical outcomes were obtained pre- and post-operatively, with results reported at the 1­year follow-up. Outcome measures included the Questionnaire for persons with a Trans-Femoral Amputation (Q-TFA), Short Form Health Survey 36 (SF-36), Six-Minute Walk Test (6MWT), and Timed Up and Go (TUG). Adverse events were also recorded. RESULTS: Compared to the mean pre-operative values obtained while patients were using socket prostheses or were wheelchair-bound, the mean post-operative values for all four validated outcome measures were significantly improved. There were 15 episodes of minor infections in 12 patients, all of which responded to antibiotics. Soft tissue refashioning was performed electively on 6 patients. No other adverse events were recorded. CONCLUSIONS: The results demonstrate that osseointegration surgery using the OPL is a relatively safe and effective procedure for the reconstruction and rehabilitation of lower limb amputees.

MiR-9-5p, miR-675-5p and miR-138-5p Damages the Strontium and LRP5-Mediated Skeletal Cell Proliferation, Differentiation, and Adhesion.

This study was designed to evaluate the effects of strontium on the expression levels of microRNAs (miRNAs) and to explore their effects on skeletal cell proliferation, differentiation, adhesion, and apoptosis. The targets of these miRNAs were also studied. Molecular cloning, cell proliferation assay, cell apoptosis assay, quantitative real-time PCR, and luciferase reporter assay were used. Strontium altered the expression levels of miRNAs in vitro and in vivo. miR-9-5p, miR-675-5p, and miR-138-5p impaired skeletal cell proliferation, cell differentiation and cell adhesion. miR-9-5p and miR-675-5p induced MC3T3-E1 cell apoptosis more specifically than miR-138-5p. miR-9-5p, miR-675-5p, and miR-138-5p targeted glycogen synthase kinase 3 ß (GSK3ß), ATPase Aminophospholipid Transporter Class I Type 8A Member 2 (ATP8A2), and Eukaryotic Translation Initiation Factor 4E Binding Protein 1 (EIF4EBP1), respectively. Low-density lipoprotein receptor-related protein 5 (LRP5) played a positive role in skeletal development. miR-9-5p, miR-675-5p, and miR-138-5p damage strontium and LRP5-mediated skeletal cell proliferation, differentiation, and adhesion, and induce cell apoptosis by targeting GSK3ß, ATP8A2, and EIF4EBP1, respectively.

Mesenchymal stem cells reduce intervertebral disc fibrosis and facilitate repair.

Intervertebral disc degeneration is associated with back pain and radiculopathy which, being a leading cause of disability, seriously affects the quality of life and presents a hefty burden to society. There is no effective intervention for the disease and the etiology remains unclear. Here, we show that disc degeneration exhibits features of fibrosis in humans and confirmed this in a puncture-induced disc degeneration (PDD) model in rabbit. Implantation of bone marrow-derived mesenchymal stem cells (MSCs) to PDD discs can inhibit fibrosis in the nucleus pulposus with effective preservation of mechanical properties and overall spinal function. We showed that the presence of MSCs can suppress abnormal deposition of collagen I in the nucleus pulposus, modulating profibrotic mediators MMP12 and HSP47, thus reducing collagen aggregation and maintaining proper fibrillar properties and function. As collagen fibrils can regulate progenitor cell activities, our finding provides new insight to the limited self-repair capability of the intervertebral disc and importantly the mechanism by which MSCs may potentiate tissue regeneration through regulating collagen fibrillogenesis in the context of fibrotic diseases.

The relationship between self-reported sleep disturbance and polysomnography in individuals with traumatic brain injury.

PRIMARY OBJECTIVE: To characterize sleep architecture and self-reported sleep quality, fatigue and daytime sleepiness in individuals with TBI. Possible relationships between sleep architecture and self-reported sleep quality, fatigue and daytime sleepiness were examined. METHODS: Forty-four community-dwelling adults with TBI completed the Pittsburgh Sleep Quality Index (PSQI), Multidimensional Assessment of Fatigue (MAF) and Epworth Sleepiness Scale (ESS). They underwent two nights of in-laboratory nocturnal polysomnography (NPSG). Pearson product-moment correlation coefficients and hierarchical linear regression was used to analyse the data. RESULTS: Based on the PSQI cut-off score of ≥ 10, 22 participants were characterized as poor sleepers. Twenty-seven participants met criteria for clinically significant fatigue as measured by the GFI of the MAF. Fourteen participants met criteria for excessive daytime sleepiness as measured by the ESS. Poor sleep quality was associated with poor sleep efficiency, short duration of stage 2 sleep and long duration of rapid eye movement sleep. There was little-to-no association between high levels of fatigue or daytime sleepiness with NPSG sleep parameters. CONCLUSIONS: A high proportion of the sample endorsed poor sleep quality, fatigue and daytime sleepiness. Those who reported poorer sleep quality evidenced a shorter proportion of time spent in stage 2 sleep. These findings suggest that disruptions in stage 2 sleep might underlie the symptoms of sleep disturbance experienced following TBI.