Varying diffusion time to discriminate between simulated skeletal muscle injury models using stimulated echo diffusion tensor imaging.

PURPOSE: Evaluate the relationship between muscle microstructure, diffusion time (Δ), and the diffusion tensor (DT) to identify the optimal Δ where changes in muscle fiber size may be detected. METHODS: The DT was simulated in models with histology informed geometry over a range of Δ with a stimulated echo DT imaging (DTI) sequence using the numerical simulation application DifSim. The difference in the DT at each Δ between healthy and injured skeletal muscle models was calculated, to identify the optimal Δ at which changes in muscle fiber size may be detected. The random permeable barrier model (RPBM) was used to estimate muscle microstructure from the simulated DT measurements, which were compared to the ground truth. RESULTS: Across all models, fractional anisotropy provided greater contrast between injured and control models than diffusivity measurements. Compared to control models, in atrophic injury models, the greatest difference in the DT was found between 90 ms and 250 ms. In models with acute edema, the contrast between injured and control muscle increased with increasing diffusion time, although these models had smaller mean fiber areas. RPBM systematically underestimated fiber size but accurately estimated surface area-to-volume ratio of simulated models. CONCLUSION: These findings may better inform pulse sequence parameter selection when performing DTI experiments in vivo. If only a single diffusion experiment can be performed, the selected Δ should be ~170 ms to maximize the ability to discriminate between different injury models. Ideally several diffusion times between 90 ms and 500 ms should be sampled in order to maximize diffusion contrast, particularly when the disease process is unknown.

Intravoxel incoherent motion imaging predicts exercise-based rehabilitation response in individuals with low back pain.

Exercises to strengthen and stabilize the trunk musculature are a common conservative treatment strategy for low back pain (LBP), despite the possible presence of impairments in muscle activation in this population. Intravoxel incoherent motion (IVIM) MRI permits evaluation of activation-induced blood flow through diffusion-weighted images that are sensitized to microvascular blood flow. In the current study we aimed to evaluate IVIM signal changes after exercise in patients with LBP compared with pain-free healthy controls and determine if these changes were related to reductions in disability with a 12-week rehabilitation program. We hypothesize that the magnitude of changes in IVIM parameters in the lumbar extensor muscles will be smaller in patients with LBP compared with those without LBP, and that these magnitudes will be correlated with responsiveness to a 12-week, resistance-based exercise program. IVIM MR data for molecular diffusion (D), blood flow pseudodiffusion (D*) and perfusion fraction (f) were collected before and immediately after an ~ 3-min session of high-intensity lumbar extension resistance exercise in 16 healthy participants and 17 participants with LBP. Improvements in LBP-related disability after the 12-week, machine-based, high-intensity exercise rehabilitation program were measured in the LBP group. We observed a significant increase in all IVIM parameters (f, D*, D) in response to exercise (p < 0.0001) and an interaction of group-by-time for D (p = 0.016). Thresholds were identified using receiver operating characteristic (ROC) curves for diffusion and pseudodiffusion coefficients, which predicted a reduction in LBP-related disability in response to the 12-week, exercise-based rehabilitation program. Exercise was associated with an increase in (f), capillary blood flow-based pseudodiffusion (D*) and diffusion coefficient (D), regardless of the presence of LBP. Additionally, subgroup analysis identified patients who were not responsive to the acute exercise session, for whom, based on ROC analysis, there was no clinically significant change in disability following the 12-week program.

3D printing of a biocompatible double network elastomer with digital control of mechanical properties.

The majority of 3D-printed biodegradable biomaterials are brittle, limiting their potential application to compliant tissues. Poly (glycerol sebacate) acrylate (PGSA) is a synthetic biodegradable and biocompatible elastomer, compatible with light-based 3D printing. In this work we employed digital-light-processing (DLP)-based 3D printing to create a complex PGSA network structure. Nature-inspired double network (DN) structures with two geometrically interconnected segments with different mechanical properties were printed from the same material in a single shot. Such capability has not been demonstrated by any other fabrication technique. The biocompatibility of PGSA after 3D printing was confirmed via cell-viability analysis. We used a finite element analysis (FEA) model to predict the failure of the DN structure under uniaxial tension. FEA confirmed the soft segments act as sacrificial elements while the hard segments retain structural integrity. The simulation demonstrated that the DN design absorbs 100% more energy before rupture than the network structure made by single exposure condition (SN), doubling the toughness of the overall structure. Using the FEA-informed design, a new DN structure was printed and the FEA predicted tensile test results agreed with tensile testing of the printed structure. This work demonstrated how geometrically-optimized material design can be easily and rapidly achieved by using DLP-based 3D printing, where well-defined patterns of different stiffnesses can be simultaneously formed using the same elastic biomaterial, and overall mechanical properties can be specifically optimized for different biomedical applications.

Regional differences between superficial and deep lumbar multifidus in patients with chronic lumbar spine pathology.

BACKGROUND: Due to its unique arrangement, the deep and superficial fibers of the multifidus may have differential roles for maintaining spine stabilization and lumbar posture; the superficial multifidus is responsible for lumbar extension and the deep multifidus for intersegmental stability. In patients with chronic lumbar spine pathology, muscle activation patterns have been shown to be attenuated or delayed in the deep, but not superficial, multifidus. This has been interpreted as pain differentially influencing the deep region. However, it is unclear if degenerative changes affecting the composition and function of the multifidus differs between the superficial and deep regions, an alternative explanation for these electrophysiological changes. Therefore, the goal of this study was to investigate macrostructural and microstructural differences between the superficial and deep regions of the multifidus muscle in patients with lumbar spine pathology. METHODS: In 16 patients undergoing lumbar spinal surgery for degenerative conditions, multifidus biopsies were acquired at two distinct locations: 1) the most superficial portion of muscle adjacent to the spinous process and 2) approximately 1 cm lateral to the spinous process and deeper at the spinolaminar border of the affected vertebral level. Structural features related to muscle function were histologically compared between these superficial and deep regions, including tissue composition, fat fraction, fiber cross sectional area, fiber type, regeneration, degeneration, vascularity and inflammation. RESULTS: No significant differences in fat signal fraction, muscle area, fiber cross sectional area, muscle regeneration, muscle degeneration, or vascularization were found between the superficial and deep regions of the multifidus. Total collagen content between the two regions was the same. However, the superficial region of the multifidus was found to have less loose and more dense collagen than the deep region. CONCLUSIONS: The results of our study did not support that the deep region of the multifidus is more degenerated in patients with lumbar spine pathology, as gross degenerative changes in muscle microstructure and macrostructure were the same in the superficial and deep regions of the multifidus. In these patients, the multifidus is not protected in order to maintain mobility and structural stability of the spine.

The effect of high-intensity resistance exercise on lumbar musculature in patients with low back pain: a preliminary study.

BACKGROUND: Muscle atrophy and fatty infiltration of the lumbar extensors is associated with LBP. Exercise-based rehabilitation targets strengthening these muscles, but few studies show consistent changes in muscle quality with standard-of-care rehabilitation. The goal of this study was to assess the effect of high-intensity resistance exercise on lumbar extensor muscle size (cross sectional area) and quality (fat fraction) in individuals with low back pain (LBP). METHODS: Fourteen patients with LBP were recruited from a local rehabilitation clinic. Patients underwent MRI scanning before and after a standardized 10-week high-intensity machine-based, resistance exercise program. Patient pain, disability, anxiety/depression, satisfaction, strength, and range of motion was compared pre- and post-rehabilitation using analysis of covariance (covariates: age, gender). Exercise-induced changes in MRI, and patient functional outcome measures were correlated using Pearson's correlation test. RESULTS: No significant differences were found in muscle size or fatty infiltration of the lumbar extensors over the course of rehabilitation (p > 0.31). However, patients reported reduced pain (p = 0.002) and were stronger (p = 0.03) at the conclusion of the program. Improvements in muscle size and quality for both multifidus and erector spinae correlated with improvements in disability, anxiety/depression, and strength. CONCLUSION: While average muscle size and fatty infiltration levels did not change with high-intensity exercise, the results suggest that a subgroup of patients who demonstrate improvements in muscle health demonstrate the largest functional improvements. Future research is needed to identify which patients are most likely to respond to this type of treatment.

Relationships between tissue microstructure and the diffusion tensor in simulated skeletal muscle.

PURPOSE: To establish a series of relationships defining how muscle microstructure and diffusion tensor imaging (DTI) are related. METHODS: The relationship among key microstructural features of skeletal muscle (fiber size, fibrosis, edema, and permeability) and the diffusion tensor were systematically simulated over physiologically relevant dimensions individually, and in combination, using a numerical simulation application. Stepwise multiple regression was used to identify which microstructural features of muscle significantly predict the diffusion tensor using single-echo and multi-echo DTI pulse sequences. Simulations were also performed in models with histology-informed geometry to investigate the relationship between fiber size and the diffusion tensor in models with real muscle geometry. RESULTS: Fiber size is the strongest predictor of λ2, λ3, mean diffusivity, and fractional anisotropy in skeletal muscle, accounting for approximately 40% of the variance in the diffusion model when calculated with single-echo DTI. This increased to approximately 70% when diffusion measures were calculated from the short T2 component of the multi-echo DTI sequence. This nonlinear relationship begins to plateau in fibers with greater than 60-µm diameter. CONCLUSIONS: As the normal fiber size of a human muscle fiber is 40 to 60 µm, this suggests that DTI is a sensitive tool to monitor muscle atrophy, but may be limited in measurements of muscle with larger fibers. Magn Reson Med 80:317-329, 2018. © 2017 International Society for Magnetic Resonance in Medicine.

Methodological considerations in region of interest definitions for paraspinal muscles in axial MRIs of the lumbar spine.

BACKGROUND: Magnetic Resonance Imaging (MRI) is commonly used to assess the health of the lumbar spine and supporting structures. Studies have suggested that fatty infiltration of the posterior lumbar muscles is important in predicting responses to treatment for low back pain. However, methodological differences exist in defining the region of interest (ROI) of a muscle, which limits the ability to compare data between studies. The purpose of this study was to determine reliability and systematic differences within and between two commonly utilized methodologies for ROI definitions of lumbar paraspinal muscle. METHODS: T2-weighted MRIs of the mid-L4 vertebrae from 37 patients with low back pain who were scheduled for lumbar spine surgery were included from a hospital database. Fatty infiltration for these patients ranged from low to high, based on Kjaer criteria. Two methods were used to define ROI: 1) segmentation of the multifidus and erector spinae based on fascial planes including epimuscular fat, and 2) segmentation of the multifidus and erector spinae based on visible muscle boundaries, which did not include epimuscular fat. Total cross sectional area (tCSA), fat signal fraction (FSF), muscle cross sectional area, and fat cross sectional area were measured. Degree of agreement between raters for each parameter was assessed using intra-class correlation coefficients (ICC) and area fraction of overlapping voxels. RESULTS: Excellent inter-rater agreement (ICC > 0.75) was observed for all measures for both methods. There was no significant difference between area fraction overlap of ROIs between methods. Method 1 demonstrated a greater tCSA for both the erector spinae (14-15%, p < 0.001) and multifidus (4%, p < 0.016) but a greater FSF only for the erector spinae (11-13%, p < 0.001). CONCLUSION: The two methods of defining lumbar spine muscle ROIs demonstrated excellent inter-rater reliability, although significant differences exist as method 1 showed larger CSA and FSF values compared to method 2. The results of this study confirm the validity of using either method to measure lumbar paraspinal musculature, and that method should be selected based on the primary outcome variables of interest.

Evidence for α-synuclein prions causing multiple system atrophy in humans with parkinsonism.

Prions are proteins that adopt alternative conformations that become self-propagating; the PrP(Sc) prion causes the rare human disorder Creutzfeldt-Jakob disease (CJD). We report here that multiple system atrophy (MSA) is caused by a different human prion composed of the α-synuclein protein. MSA is a slowly evolving disorder characterized by progressive loss of autonomic nervous system function and often signs of parkinsonism; the neuropathological hallmark of MSA is glial cytoplasmic inclusions consisting of filaments of α-synuclein. To determine whether human α-synuclein forms prions, we examined 14 human brain homogenates for transmission to cultured human embryonic kidney (HEK) cells expressing full-length, mutant human α-synuclein fused to yellow fluorescent protein (α-syn140*A53T-YFP) and TgM83(+/-) mice expressing α-synuclein (A53T). The TgM83(+/-) mice that were hemizygous for the mutant transgene did not develop spontaneous illness; in contrast, the TgM83(+/+) mice that were homozygous developed neurological dysfunction. Brain extracts from 14 MSA cases all transmitted neurodegeneration to TgM83(+/-) mice after incubation periods of ∼120 d, which was accompanied by deposition of α-synuclein within neuronal cell bodies and axons. All of the MSA extracts also induced aggregation of α-syn*A53T-YFP in cultured cells, whereas none of six Parkinson's disease (PD) extracts or a control sample did so. Our findings argue that MSA is caused by a unique strain of α-synuclein prions, which is different from the putative prions causing PD and from those causing spontaneous neurodegeneration in TgM83(+/+) mice. Remarkably, α-synuclein is the first new human prion to be identified, to our knowledge, since the discovery a half century ago that CJD was transmissible.

The effect of training on lumbar spine posture and intervertebral disc degeneration in active-duty Marines.

Military training aims to improve load carriage performance and reduce risk of injuries. Data describing the lumbar spine (LS) postural response to load carriage throughout training are limited. We hypothesised that training would reduce the LS postural response to load. The LS posture of 27 Marines was measured from upright MR images: with and without load (22.6 kg) at the beginning, middle, and end of School of Infantry (SOI) training. Disc degeneration was graded at L5-S1. No changes in posture and disc degeneration were found throughout training. During load carriage the LS became less lordotic and the sacrum more horizontal. Marines with disc degeneration had larger sacral postural perturbations in response to load. Our findings suggest that the postural response to load is defined more by the task needs than by the physical condition of the Marine. Practitioner Summary: The effect of military training on lumbar spine posture is unknown. The lumbar posture of 27 Marines was measured from upright MR images, with and without load throughout infantry training. No changes in posture or IVD degeneration were found across training. Marines with degeneration at the L5-S1 level had larger sacral postural perturbations in response to load.

Optimization of Aryl Amides that Extend Survival in Prion-Infected Mice.

Developing therapeutics for neurodegenerative diseases (NDs) prevalent in the aging population remains a daunting challenge. With the growing understanding that many NDs progress by conformational self-templating of specific proteins, the prototypical prion diseases offer a platform for ND drug discovery. We evaluated high-throughput screening hits with the aryl amide scaffold and explored the structure-activity relationships around three series differing in their N-aryl core: benzoxazole, benzothiazole, and cyano. Potent anti-prion compounds were advanced to pharmacokinetic studies, and the resulting brain-penetrant leads from each series, together with a related N-aryl piperazine lead, were escalated to long-term dosing and efficacy studies. Compounds from each of the four series doubled the survival of mice infected with a mouse-passaged prion strain. Treatment with aryl amides altered prion strain properties, as evidenced by the distinct patterns of neuropathological deposition of prion protein and associated astrocytic gliosis in the brain; however, none of the aryl amide compounds resulted in drug-resistant prion strains, in contrast to previous studies on compounds with the 2-aminothiazole (2-AMT) scaffold. As seen with 2-AMTs and other effective anti-prion compounds reported to date, the novel aryl amides reported here were ineffective in prolonging the survival of transgenic mice infected with human prions. Most encouraging is our discovery that aryl amides show that the development of drug resistance is not an inevitable consequence of efficacious anti-prion therapeutics.

Drug resistance confounding prion therapeutics.

There is not a single pharmaceutical that halts or even slows any neurodegenerative disease. Mounting evidence shows that prions cause many neurodegenerative diseases, and arguably, scrapie and Creutzfeldt-Jakob disease prions represent the best therapeutic targets. We report here that the previously identified 2-aminothiazoles IND24 and IND81 doubled the survival times of scrapie-infected, wild-type mice. However, mice infected with Rocky Mountain Laboratory (RML) prions, a scrapie-derived strain, and treated with IND24 eventually exhibited neurological dysfunction and died. We serially passaged their brain homogenates in mice and cultured cells. We found that the prion strain isolated from IND24-treated mice, designated RML[IND24], emerged during a single passage in treated mice. Although RML prions infect both the N2a and CAD5 cell lines, RML[IND24] prions could only infect CAD5 cells. When passaged in CAD5 cells, the prions remained resistant to high concentrations of IND24. However, one passage of RML[IND24] prions in untreated mice restored susceptibility to IND24 in CAD5 cells. Although IND24 treatment extended the lives of mice propagating different prion strains, including RML, another scrapie-derived prion strain ME7, and chronic wasting disease, it was ineffective in slowing propagation of Creutzfeldt-Jakob disease prions in transgenic mice. Our studies demonstrate that prion strains can acquire resistance upon exposure to IND24 that is lost upon passage in mice in the absence of IND24. These data suggest that monotherapy can select for resistance, thus intermittent therapy with mixtures of antiprion compounds may be required to slow or stop neurodegeneration.

Different 2-Aminothiazole Therapeutics Produce Distinct Patterns of Scrapie Prion Neuropathology in Mouse Brains.

Because no drug exists that halts or even slows any neurodegenerative disease, developing effective therapeutics for any prion disorder is urgent. We recently reported two compounds (IND24 and IND81) with the 2-aminothiazole (2-AMT) chemical scaffold that almost doubled the incubation times in scrapie prion-infected, wild-type (wt) FVB mice when given in a liquid diet. Remarkably, oral prophylactic treatment with IND24 beginning 14 days prior to intracerebral prion inoculation extended survival from ∼120 days to over 450 days. In addition to IND24, we evaluated the pharmacokinetics and efficacy of five additional 2-AMTs; one was not followed further because its brain penetration was poor. Of the remaining four new 2-AMTs, IND114338 doubled and IND125 tripled the incubation times of RML-inoculated wt and Tg4053 mice overexpressing wt mouse prion protein (PrP), respectively. Neuropathological examination of the brains from untreated controls showed a widespread deposition of self-propagating, ß-sheet-rich "scrapie" isoform (PrP(Sc)) prions accompanied by a profound astrocytic gliosis. In contrast, mice treated with 2-AMTs had lower levels of PrP(Sc) and associated astrocytic gliosis, with each compound resulting in a distinct pattern of deposition. Notably, IND125 prevented both PrP(Sc) accumulation and astrocytic gliosis in the cerebrum. Progressive central nervous system dysfunction in the IND125-treated mice was presumably due to the PrP(Sc) that accumulated in their brainstems. Disappointingly, none of the four new 2-AMTs prolonged the lives of mice expressing a chimeric human/mouse PrP transgene inoculated with Creutzfeldt-Jakob disease prions.

Pathway connectivity and signaling coordination in the yeast stress-activated signaling network.

Stressed cells coordinate a multi-faceted response spanning many levels of physiology. Yet knowledge of the complete stress-activated regulatory network as well as design principles for signal integration remains incomplete. We developed an experimental and computational approach to integrate available protein interaction data with gene fitness contributions, mutant transcriptome profiles, and phospho-proteome changes in cells responding to salt stress, to infer the salt-responsive signaling network in yeast. The inferred subnetwork presented many novel predictions by implicating new regulators, uncovering unrecognized crosstalk between known pathways, and pointing to previously unknown 'hubs' of signal integration. We exploited these predictions to show that Cdc14 phosphatase is a central hub in the network and that modification of RNA polymerase II coordinates induction of stress-defense genes with reduction of growth-related transcripts. We find that the orthologous human network is enriched for cancer-causing genes, underscoring the importance of the subnetwork's predictions in understanding stress biology.

An Endplate-Based Joint Coordinate System for Measuring Kinematics in Normal and Abnormally-Shaped Lumbar Vertebrae.

Vertebral level-dependent, angular, and linear translations of the spine have been measured in 2D and 3D using several imaging methods to quantify postural changes due to loading conditions and tasks. Here, we propose and validate a semiautomated method for measuring lumbar intervertebral angles and translations from upright MRI images using an endplate-based, joint coordinate system (JCS). This method was validated using 3D printed structures, representing intervertebral discs (IVD) at predetermined angles and heights, which were positioned between adjacent cadaveric vertebrae as a gold standard. Excellent agreement between our measurements and the gold standard was found for intervertebral angles in all anatomical planes (ICC > .997) and intervertebral distance measurements (ICC > .949). The proposed endplate-based JCS was compared with the vertebral body-based JCS proposed by the International Society of Biomechanics (ISB) using the 3D printed structures placed between 3 adjacent vertebrae from a cadaver with scoliosis. The endplate-based method was found to have better agreement with angles in the sagittal plane (ICC = 0.985) compared with the vertebral body-based method (ICC = .280). Thus, this method is accurate for measuring 3D intervertebral angles in the healthy and diseased lumbar spine.

Multiple means to the same end: the genetic basis of acquired stress resistance in yeast.

In nature, stressful environments often occur in combination or close succession, and thus the ability to prepare for impending stress likely provides a significant fitness advantage. Organisms exposed to a mild dose of stress can become tolerant to what would otherwise be a lethal dose of subsequent stress; however, the mechanism of this acquired stress tolerance is poorly understood. To explore this, we exposed the yeast gene-deletion libraries, which interrogate all essential and non-essential genes, to successive stress treatments and identified genes necessary for acquiring subsequent stress resistance. Cells were exposed to one of three different mild stress pretreatments (salt, DTT, or heat shock) and then challenged with a severe dose of hydrogen peroxide (H(2)O(2)). Surprisingly, there was little overlap in the genes required for acquisition of H(2)O(2) tolerance after different mild-stress pretreatments, revealing distinct mechanisms of surviving H(2)O(2) in each case. Integrative network analysis of these results with respect to protein-protein interactions, synthetic-genetic interactions, and functional annotations identified many processes not previously linked to H(2)O(2) tolerance. We tested and present several models that explain the lack of overlap in genes required for H(2)O(2) tolerance after each of the three pretreatments. Together, this work shows that acquired tolerance to the same severe stress occurs by different mechanisms depending on prior cellular experiences, underscoring the context-dependent nature of stress tolerance.

Impact of Deltoid Computer Tomography Image Data on the Accuracy of Machine Learning Predictions of Clinical Outcomes after Anatomic and Reverse Total Shoulder Arthroplasty.

Background: Despite the importance of the deltoid to shoulder biomechanics, very few studies have quantified the three-dimensional shape, size, or quality of the deltoid muscle, and no studies have correlated these measurements to clinical outcomes after anatomic (aTSA) and/or reverse (rTSA) total shoulder arthroplasty in any statistically/scientifically relevant manner. Methods: Preoperative computer tomography (CT) images from 1057 patients (585 female, 469 male; 799 primary rTSA and 258 primary aTSA) of a single platform shoulder arthroplasty prosthesis (Equinoxe; Exactech, Inc., Gainesville, FL) were analyzed in this study. A machine learning (ML) framework was used to segment the deltoid muscle for 1057 patients and quantify 15 different muscle characteristics, including volumetric (size, shape, etc.) and intensity-based Hounsfield (HU) measurements. These deltoid measurements were correlated to postoperative clinical outcomes and utilized as inputs to train/test ML algorithms used to predict postoperative outcomes at multiple postoperative timepoints (1 year, 2-3 years, and 3-5 years) for aTSA and rTSA. Results: Numerous deltoid muscle measurements were demonstrated to significantly vary with age, gender, prosthesis type, and CT image kernel; notably, normalized deltoid volume and deltoid fatty infiltration were demonstrated to be relevant to preoperative and postoperative clinical outcomes after aTSA and rTSA. Incorporating deltoid image data into the ML models improved clinical outcome prediction accuracy relative to ML algorithms without image data, particularly for the prediction of abduction and forward elevation after aTSA and rTSA. Analyzing ML feature importance facilitated rank-ordering of the deltoid image measurements relevant to aTSA and rTSA clinical outcomes. Specifically, we identified that deltoid shape flatness, normalized deltoid volume, deltoid voxel skewness, and deltoid shape sphericity were the most predictive image-based features used to predict clinical outcomes after aTSA and rTSA. Many of these deltoid measurements were found to be more predictive of aTSA and rTSA postoperative outcomes than patient demographic data, comorbidity data, and diagnosis data. Conclusions: While future work is required to further refine the ML models, which include additional shoulder muscles, like the rotator cuff, our results show promise that the developed ML framework can be used to evolve traditional CT-based preoperative planning software into an evidence-based ML clinical decision support tool.

Paraspinal muscles in individuals undergoing surgery for lumbar spine pathology lack a myogenic response to an acute bout of resistance exercise.

Background: Lumbar spine pathology (LSP) is a common source of low back or leg pain, and paraspinal muscle in these patients demonstrates fatty and fibrotic infiltration, and cellular degeneration that do not reverse with exercise-based rehabilitation. However, it is unclear of this lack of response is due to insufficient exercise stimulus, or an inability to mount a growth response. The purpose of this study was to compare paraspinal muscle gene expression between individuals with LSP who do and do not undergo an acute bout of resistance exercise. Methods: Paraspinal muscle biopsies were obtained from 64 individuals with LSP undergoing spinal surgery. Eight participants performed an acute bout of machine-based lumbar extension resistance exercise preoperatively. Gene expression for 42 genes associated with adipogenic/metabolic, atrophic, fibrogenic, inflammatory, and myogenic pathways was measured, and differential expression between exercised and non-exercised groups was evaluated for (a) the full cohort, and (b) an age, gender, acuity, and etiology matched sub-cohort. Principal components analyses were used to identify gene expression clustering across clinical phenotypes. Results: The exercised cohort demonstrated upregulation of inflammatory gene IL1B, inhibition of extracellular matrix components (increased MMP3&9, decreased TIMP1&3, COL1A1) and metabolic/adipogenic genes (FABP4, PPARD, WNT10B), and downregulation of myogenic (MYOD, ANKRD2B) and atrophic (FOXO3) genes compared to the non-exercised cohort, with similar patterns in the matched sub-analysis. There were no clinical phenotypes significantly associated with gene expression profiles. Conclusion: An acute bout of moderate-high intensity resistance exercise did not result in upregulation of myogenic genes in individuals with LSP. The response was characterized by mixed metabolic and fibrotic gene expression, upregulation of inflammation, and downregulation of myogenesis.

Effect of specimen-specific anisotropic material properties in quantitative computed tomography-based finite element analysis of the vertebra.

Intra- and inter-specimen variations in trabecular anisotropy are often ignored in quantitative computed tomography (QCT)-based finite element (FE) models of the vertebra. The material properties are typically estimated solely from local variations in bone mineral density (BMD), and a fixed representation of elastic anisotropy ("generic anisotropy") is assumed. This study evaluated the effect of incorporating specimen-specific, trabecular anisotropy on QCT-based FE predictions of vertebral stiffness and deformation patterns. Orthotropic material properties estimated from microcomputed tomography data ("specimen-specific anisotropy"), were assigned to a large, columnar region of the L1 centrum (n = 12), and generic-anisotropic material properties were assigned to the remainder of the vertebral body. Results were compared to FE analyses in which generic-anisotropic properties were used throughout. FE analyses were also performed on only the columnar regions. For the columnar regions, the axial stiffnesses obtained from the two categories of material properties were uncorrelated with each other (p = 0.604), and the distributions of minimum principal strain were distinctly different (p ≤ 0.022). In contrast, for the whole vertebral bodies in both axial and flexural loading, the stiffnesses obtained using the two categories of material properties were highly correlated (R2 > 0.82, p < 0.001) with, and were no different (p > 0.359) from, each other. Only moderate variations in strain distributions were observed between the two categories of material properties. The contrasting results for the columns versus vertebrae indicate a large contribution of the peripheral regions of the vertebral body to the mechanical behavior of this bone. In companion analyses on the effect of the degree of anisotropy (DA), the axial stiffnesses of the trabecular column (p < 0.001) and vertebra (p = 0.007) increased with increasing DA. These findings demonstrate the need for accurate modeling of the peripheral regions of the vertebral body in analyses of the mechanical behavior of the vertebra.

Multimodal Three-Dimensional Printing for Micro-Modulation of Scaffold Stiffness Through Machine Learning.

The ability to precisely control a scaffold's microstructure and geometry with light-based three-dimensional (3D) printing has been widely demonstrated. However, the modulation of scaffold's mechanical properties through prescribed printing parameters is still underexplored. This study demonstrates a novel 3D-printing workflow to create a complex, elastomeric scaffold with precision-engineered stiffness control by utilizing machine learning. Various printing parameters, including the exposure time, light intensity, printing infill, laser pump current, and printing speed were modulated to print poly (glycerol sebacate) acrylate (PGSA) scaffolds with mechanical properties ranging from 49.3 ± 3.3 kPa to 2.8 ± 0.3 MPa. This enables flexibility in spatial stiffness modulation in addition to high-resolution scaffold fabrication. Then, a neural network-based machine learning model was developed and validated to optimize printing parameters to yield scaffolds with user-defined stiffness modulation for two different vat photopolymerization methods: a digital light processing (DLP)-based 3D printer was utilized to rapidly fabricate stiffness-modulated scaffolds with features on the hundreds of micron scale and a two-photon polymerization (2PP) 3D printer was utilized to print fine structures on the submicron scale. A novel 3D-printing workflow was designed to utilize both DLP-based and 2PP 3D printers to create multiscale scaffolds with precision-tuned stiffness control over both gross and fine geometric features. The described workflow can be used to fabricate scaffolds for a variety of tissue engineering applications, specifically for interfacial tissue engineering for which adjacent tissues possess heterogeneous mechanical properties (e.g., muscle-tendon).

High cell density and high-resolution 3D bioprinting for fabricating vascularized tissues.

Three-dimensional (3D) bioprinting techniques have emerged as the most popular methods to fabricate 3D-engineered tissues; however, there are challenges in simultaneously satisfying the requirements of high cell density (HCD), high cell viability, and fine fabrication resolution. In particular, bioprinting resolution of digital light processing-based 3D bioprinting suffers with increasing bioink cell density due to light scattering. We developed a novel approach to mitigate this scattering-induced deterioration of bioprinting resolution. The inclusion of iodixanol in the bioink enables a 10-fold reduction in light scattering and a substantial improvement in fabrication resolution for bioinks with an HCD. Fifty-micrometer fabrication resolution was achieved for a bioink with 0.1 billion per milliliter cell density. To showcase the potential application in tissue/organ 3D bioprinting, HCD thick tissues with fine vascular networks were fabricated. The tissues were viable in a perfusion culture system, with endothelialization and angiogenesis observed after 14 days of culture.