T1Gd is reduced in bone marrow lesions overlying cartilage in the hip.

OBJECTIVE: Bone Marrow Lesions (BMLs) are areas in bone with high fluid signal on MRI associated with painful and progressive OA. While cartilage near BMLs in the knee has been shown to be degenerated, this relationship has not been investigated in the hip. RESEARCH QUESTION: is T1Gd lower in areas of cartilage overlying BMLs in the hip? DESIGN: 128 participants were recruited from a population-based study of hip pain in 20-49-year-olds. Proton-density weighted fat-suppressed and delayed Gadolinium Enhanced MR Imaging of Cartilage (dGEMRIC) images were acquired to locate BMLs and quantify hip cartilage health. BML and cartilage images were registered and cartilage was separated into BML overlying and surrounding regions. Mean T1Gd was measured in 32 participants with BMLs in both cartilage regions and in matched regions in 32 age- and sex-matched controls. Mean T1Gd in the overlying cartilage was compared using linear mixed-effects models between BML and control groups for acetabular and femoral BMLs, and between cystic and non-cystic BML groups. RESULTS: Mean T1Gd of overlying cartilage was lower in the BML group compared to the control group (acetabular: -105 ms; 95% CI: -175, -35; femoral: -8 ms; 95% CI: -141, 124). Mean T1Gd in overlying cartilage was lower in cystic compared to non-cystic BML subjects, but the confidence interval is too large to provide certainty in this difference (-3 [95% CI: -126, 121]). CONCLUSIONS: T1Gd is reduced in overlying cartilage in hips from a population-based sample of adults aged 20-49, which suggests BMLs are associated with local cartilage degeneration in hips.

Delayed Gadolinium-Enhanced Magnetic Resonance Imaging of Cartilage Values in Hips With Bone Marrow Lesions.

OBJECTIVE: Bone marrow lesions (BMLs) are associated with painful and progressive osteoarthritis (OA). Quantitative magnetic resonance imaging (MRI) has been used to study early cartilage degeneration in knees with BML, but similar work has not been done in hips. The purpose of this study was to compare mean delayed gadolinium-enhanced MRI of cartilage (dGEMRIC) relaxation values (T1Gd) in hips with BML to hips without BML in a population-based study. Reduced T1Gd suggests depleted glycosaminoglycan. Our hypothesis was that mean T1Gd is lower in hips with BML compared to hips without BML. METHODS: Study participants (n = 128) were recruited from a cross-sectional population-based study of people ages 20-49 years with and without hip pain. dGEMRIC and proton density (PD)-weighted MRI scans of 1 hip from each participant were used for this analysis. BMLs were identified from PD-weighted fat-suppressed images. We applied a sampling-weighted linear regression model to determine the association of the presence of BMLs with mean cartilage T1Gd (significance: P < 0.05). The model was adjusted for age, sex, body mass index (BMI), hip pain, cam/pincer deformity, and physical activity. RESULTS: Thirty-two (25%) of the 128 participants had at least 1 BML. Subjects with at least 1 BML, compared to those without, had similar weighted characteristics of age, BMI, physical activity levels, and frequency of hip pain. Mean T1Gd was 75.25 msec lower (95% confidence interval -149.69, -0.81; P = 0.048) (9%) in the BML compared to the no-BML group. CONCLUSION: Our results suggest that hips with BMLs are associated with hip cartilage degeneration early in the OA disease process.

Effect of posture and femoral neck osteochondroplasty on femur-acetabulum clearance in patients with cam-type femoroacetabular impingement.

It is not clear whether femoral neck osteochondroplasty achieves its objective of increasing femoroacetabular clearance. We used an upright open magnetic resonance imaging scanner to image the hip joint in multiple postures to explore the effect of posture and femoral neck osteochondroplasty on femur-acetabulum clearance in patients with cam-type femoroacetabular impingement. We recruited 13 consecutive patients scheduled to undergo arthroscopic femoral neck osteochondroplasty and completed assessments on 10 patients. We scanned each subject before surgery and at 6 months post-op in supine and 3 other physiological postures: supine 90° flexion with adduction and internal rotation (FADIR), sitting deep (maximal flexion with internal rotation and adduction), and sitting crossed leg (maximal adduction with flexion and internal rotation). We measured the alpha angle, which describes the severity of cam deformity, and the beta angle, which defines joint clearance. We also evaluated hip flexion, internal rotation, and adduction before and after surgery. Femoral neck osteochondroplasty significantly decreased alpha angle by 23.9° ± 4.6° (p = 0.001) and increased beta angle across all postures by 28.1° ± 6.3° (p = 0.002). An increase in beta angle represented a decreased chance of impingement. Femoral neck osteochondroplasty significantly increased flexion by an average of 8.6° in the sitting deep posture after surgery (p = 0.007) which might indicate an improvement of joint function. These findings lend support to the hypothesis that arthroscopic osteochondroplasty accomplishes its stated goals of increasing bone-bone clearance in the hip joint and improving joint mechanics for the static postures assessed.

Effect of positioning error on the Hilgenreiner epiphyseal angle and the head-shaft angle compared to the femoral neck-shaft angle in children with cerebral palsy.

Children with cerebral palsy (CP) often have changes in proximal femoral geometry. Neck-shaft angle (NSA), Hilgenreiner epiphyseal angle (HEA) and head-shaft angle (HSA) are used to measure these changes. The impact of femoral rotation on HEA/HSA and of ab/adduction on HEA/HSA/NSA is not well known. This study aimed to determine and compare the effect of rotation, ab/adduction and flexion/extension on HEA/HSA/NSA. Radiographic measurements from 384 patients with Gross Motor Function Classification System (GMFCS) levels I-V were utilized. NSA/HSA for affected hips were used with femoral anteversion averages to create three-dimensional models of 694 hips in children with CP. Each hip was rotated, ab/adducted and flexed/extended to simulate malpositioning. HEA/HSA/NSA of each model were measured in each joint position, and differences from correct positioning were determined. Mean HEA error at 20° of internal/external rotations were -0.60°/3.17°, respectively, with the NSA error of -6.56°/9.94° and the HSA error of -3.69°/1.21°. Each degree of ab/adduction added 1° of the HEA error, with no NSA/HSA error. NSA was most sensitive to flexion. Error for all measures increased with increasing GMFCS level. HEA/HSA were minimally impacted by rotation. NSA error was much higher than HEA/HSA in internal rotation and flexion whereas HEA was sensitive to changes in ab/adduction. Given abduction is more easily detectable on imaging than rotation, HEA may be less affected by positioning errors that are common with children with CP than NSA. HSA was least affected by position changes. HEA/HSA could be robust, complementary measures of hip deformities in children with CP.

T1ρ and T2 MRI show hip cartilage damage in adolescents with healed Legg-Calvé-Perthes disease.

Legg-Calvé-Perthes disease (LCPD) is a juvenile hip disorder associated with residual femoral head deformity, cartilage degeneration and a high risk of early onset hip osteoarthritis. Assessing management of LCPD in the healed phase requires an understanding of when and where hip cartilage damage happens. While it has been shown that cartilage is degenerated in healed LCPD hips in adults, it is not clear when this degeneration begins. Our research question was: Are the MR markers of cartilage degeneration T1ρ and T2 increased in healed LCPD hips in adolescents? Twelve adolescents [10-17 years old (mean 14); 3 female 9 male] with healed LCPD (Stulberg 2-5; 8 unilateral and 4 bilateral) and 15 age- and sex-matched controls were imaged in a 3T MRI using a T1ρ and a T2 sequence. We applied a mixed-effects model adjusted for age and nested by subject to determine the effect of Stulberg grade on overall and regional mean T1ρ and T2 values. T1ρ was significantly higher overall and in the medial region of Stulberg ≥3 hips, and in the medial region of Stulberg 2 hips than in the control group. T2 was significantly higher in the medial region of Stulberg ≥3 hips than in the control group. Our results suggest that cartilage damage in LCPD has begun by adolescence and that T1ρ can detect early changes in cartilage associated with LCPD.

Photocrosslinked Bioreducible Polymeric Nanoparticles for Enhanced Systemic siRNA Delivery as Cancer Therapy.

Clinical translation of polymer-based nanocarriers for systemic delivery of RNA has been limited due to poor colloidal stability in the blood stream and intracellular delivery of the RNA to the cytosol. To address these limitations, this study reports a new strategy incorporating photocrosslinking of bioreducible nanoparticles for improved stability extracellularly and rapid release of RNA intracellularly. In this design, the polymeric nanocarriers contain ester bonds for hydrolytic degradation and disulfide bonds for environmentally triggered small interfering RNA (siRNA) release in the cytosol. These photocrosslinked bioreducible nanoparticles (XbNPs) have a shielded surface charge, reduced adsorption of serum proteins, and enable superior siRNA-mediated knockdown in both glioma and melanoma cells in high-serum conditions compared to non-crosslinked formulations. Mechanistically, XbNPs promote cellular uptake and the presence of secondary and tertiary amines enables efficient endosomal escape. Following systemic administration, XbNPs facilitate targeting of cancer cells and tissue-mediated siRNA delivery beyond the liver, unlike conventional nanoparticle-based delivery. These attributes of XbNPs facilitate robust siRNA-mediated knockdown in vivo in melanoma tumors colonized in the lungs following systemic administration. Thus, biodegradable polymeric nanoparticles, via photocrosslinking, demonstrate extended colloidal stability and efficient delivery of RNA therapeutics under physiological conditions, and thereby potentially advance systemic delivery technologies for nucleic acid-based therapeutics.

The effects of cholesterol accumulation on Achilles tendon biomechanics: A cross-sectional study.

Familial hypercholesterolemia, a common genetic metabolic disorder characterized by high cholesterol levels, is involved in the development of atherosclerosis and other preventable diseases. Familial hypercholesterolemia can also cause tendinous abnormalities, such as thickening and xanthoma (tendon lipid accumulation) in the Achilles, which may impede tendon biomechanics. The objective of this study was to investigate the effect of cholesterol accumulation on the biomechanical performance of Achilles tendons, in vivo. 16 participants (10 men, 6 women; 37±6 years) with familial hypercholesterolemia, diagnosed with tendon xanthoma, and 16 controls (10 men, 6 women; 36±7 years) underwent Achilles biomechanical assessment. Achilles biomechanical data was obtained during preferred pace, shod, walking by analysis of lower limb kinematics and kinetics utilizing 3D motion capture and an instrumented treadmill. Gastrocnemius medialis muscle-tendon junction displacement was imaged using ultrasonography. Achilles stiffness, hysteresis, strain and force were calculated from displacement-force data acquired during loading cycles, and tested for statistical differences using one-way ANOVA. Statistical parametric mapping was used to examine group differences in temporal data. Participants with familial hypercholesterolemia displayed lower Achilles stiffness compared to the control group (familial hypercholesterolemia group: 87±20 N/mm; controls: 111±18 N/mm; p = 0.001), which appeared to be linked to Achilles loading rate rather than an increased strain (FH: 5.27±1.2%; controls: 4.95±0.9%; p = 0.413). We found different Achilles loading patterns in the familial hypercholesterolemia group, which were traced to differences in the centre of pressure progression that affected ankle moment. This finding may indicate that individuals with familial hypercholesterolemia use different Achilles loading strategies. Participants with familial hypercholesterolemia also demonstrated significantly greater Achilles hysteresis than the control group (familial hypercholesterolemia: 57.5±7.3%; controls: 43.8±10%; p<0.001), suggesting that walking may require a greater metabolic cost. Our results indicate that cholesterol accumulation could contribute to reduced Achilles function, while potentially increasing the chance of injury.

Dynamic morphometric changes in degenerative lumbar spondylolisthesis: A pilot study of upright magnetic resonance imaging.

The objectives of this study were to (a) develop a standing MRI imaging protocol, tolerable to symptomatic patients with degenerative spondylolisthesis (DLS), and (b) to evaluate the morphometric changes observed in DLS patients in both supine and standing postures. Patients with single level, Meyerding grade 1 DLS undergoing surgery at a single institution between November 2015 to May 2017 were consented. Patients were imaged in the supine and standing positions in a 0.5 T vertically open MRI scanner (MROpen, Paramed, Genoa, Italy) with sagittal and axial T2 images. The morphometric parameters measured were: cross-sectional area of the thecal sac (CSA), lateral recess height, disc height, degree of anterolisthesis, disc angle, lumbar lordosis, the presence of facet effusion and restabilization signs. Measures from both postures were compared using paired T-test. Associations of posture with the magnitude of change in the various measurements was determined using Pearson correlation or paired T-test when appropriate. All fourteen patients (mean age 64.4 years) included tolerated standing for the time required for image acquisition. All measurements with the exception of lumbar lordosis and disk height showed a statistically significant difference between the postures (p < 0.05). In the standing position, CSA and lateral recess height were reduced by 28% and 50%, respectively. There was no relationship between the change in CSA of the thecal sac and any measures. Standing images acquired in an upright MRI scanner demonstrated postural changes associated with Meyerding grade 1 DLS and images acquisition was tolerated in all patients.

In situ genetic engineering of tumors for long-lasting and systemic immunotherapy.

Cancer immunotherapy has been the subject of extensive research, but highly effective and broadly applicable methods remain elusive. Moreover, a general approach to engender endogenous patient-specific cellular therapy, without the need for a priori knowledge of tumor antigen, ex vivo cellular manipulation, or cellular manufacture, could dramatically reduce costs and broaden accessibility. Here, we describe a biotechnology based on synthetic, biodegradable nanoparticles that can genetically reprogram cancer cells and their microenvironment in situ so that the cancer cells can act as tumor-associated antigen-presenting cells (tAPCs) by inducing coexpression of a costimulatory molecule (4-1BBL) and immunostimulatory cytokine (IL-12). In B16-F10 melanoma and MC38 colorectal carcinoma mouse models, reprogramming nanoparticles in combination with checkpoint blockade significantly reduced tumor growth over time and, in some cases, cleared the tumor, leading to long-term survivors that were then resistant to the formation of new tumors upon rechallenge at a distant site. In vitro and in vivo analyses confirmed that locally delivered tAPC-reprogramming nanoparticles led to a significant cell-mediated cytotoxic immune response with systemic effects. The systemic tumor-specific and cell-mediated immunotherapy response was achieved without requiring a priori knowledge of tumor-expressed antigens and reflects the translational potential of this nanomedicine.

Nonviral polymeric nanoparticles for gene therapy in pediatric CNS malignancies.

Together, medulloblastoma (MB) and atypical teratoid/rhabdoid tumors (AT/RT) represent two of the most prevalent pediatric brain malignancies. Current treatment involves radiation, which has high risks of developmental sequelae for patients under the age of three. New safer and more effective treatment modalities are needed. Cancer gene therapy is a promising alternative, but there are challenges with using viruses in pediatric patients. We developed a library of poly(beta-amino ester) (PBAE) nanoparticles and evaluated their efficacy for plasmid delivery of a suicide gene therapy to pediatric brain cancer models-specifically herpes simplex virus type I thymidine kinase (HSVtk), which results in controlled apoptosis of transfected cells. In vivo, PBAE-HSVtk treated groups had a greater median overall survival in mice implanted with AT/RT (P = 0.0083 vs. control) and MB (P < 0.0001 vs. control). Our data provide proof of principle for using biodegradable PBAE nanoparticles as a safe and effective nanomedicine for treating pediatric CNS malignancies.

Carboxylated branched poly(ß-amino ester) nanoparticles enable robust cytosolic protein delivery and CRISPR-Cas9 gene editing.

Efficient cytosolic protein delivery is necessary to fully realize the potential of protein therapeutics. Current methods of protein delivery often suffer from low serum tolerance and limited in vivo efficacy. Here, we report the synthesis and validation of a previously unreported class of carboxylated branched poly(ß-amino ester)s that can self-assemble into nanoparticles for efficient intracellular delivery of a variety of different proteins. In vitro, nanoparticles enabled rapid cellular uptake, efficient endosomal escape, and functional cytosolic protein release into cells in media containing 10% serum. Moreover, nanoparticles encapsulating CRISPR-Cas9 ribonucleoproteins (RNPs) induced robust levels of gene knock-in (4%) and gene knockout (>75%) in several cell types. A single intracranial administration of nanoparticles delivering a low RNP dose (3.5 pmol) induced robust gene editing in mice bearing engineered orthotopic murine glioma tumors. This self-assembled polymeric nanocarrier system enables a versatile protein delivery and gene editing platform for biological research and therapeutic applications.

Patient-Specific Functional Analysis: The Key to the Next Revolution Towards the Treatment of Hip and Knee Osteoarthritis.

Lower extremity joint arthroplasty surgery remains one of the most successful interventions in orthopaedics. While improvements in patient mobility and physical functioning following surgery are well-documented, there remains significant post-operative functional deficits in many patients. This highlights a need and an opportunity towards improving functional and patient-reported outcomes of arthroplasty surgery. This review summarizes key opportunities arising from the recent 2018 Orthopaedic Research Society Meeting in New Orleans, USA. In this review, the Canadian Orthopaedic Research Society (i.e., CORS) highlights key research advances, case examples, scientific messages, and personalized medical care approaches toward improving physical functioning in our knee and hip joint arthroplasty patients. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:1754-1759, 2019.

Intraluminal Delivery of Simvastatin Attenuates Intimal Hyperplasia After Arterial Injury.

INTRODUCTION: Oral statins reduce intimal hyperplasia (IH) after arterial injury by only ∼25%. Alternative drug delivery systems have gained attention as carriers for hydrophobic drugs. We studied the effects of simvastatin (free vs hyaluronic acid-tagged polysialic acid-polycaprolactone micelles) on vascular smooth muscle cell (VSMC) migration, VSMC proliferation and intimal hyperplasia. We hypothesized both free and micelle containing simvastatin would inhibit VSMC chemotaxis and proliferation, and local statin treatment would be more effective than oral in reducing IH in rats following carotid balloon injury. METHODS: VSMCs pretreated with free simvastatin (20 minutes or 20 hours) or simvastatin-loaded micelles underwent chemotaxis and proliferation to platelet-derived growth factor. Next, rats that underwent balloon injury of the common carotid artery received statin therapy-intraluminal simvastatin-loaded micelles prior to injury, periadventitial pluronic gel following injury, or combinations of gel, micelle, and oral simvastatin. After 14 days, morphometric analysis determined the -intimal to medial ratio. Findings were compared to controls receiving oral simvastatin or no statin therapy. Statistical analysis was by analysis of variance for the in vitro experiments and a factorial general linear model for the in vivo experiments. RESULTS: The simvastatin-loaded micelles and free simvastatin inhibited VSMC chemotaxis (54%-60%). IH was induced in all injured vessels. Simvastatin in pluronic gel or micelles reduced IH compared to untreated controls (0.208 ± 0.04 or 0.160 ± 0.03 vs 0.350 ± 0.03, respectively); however, neither gel nor simvastatin-loaded micelles were superior to oral statins (0.261 ± 0.03). Addition of oral statins or combining both local therapies did not provide additional benefit. Micelles were the single greatest contributing factor in IH attenuation. CONCLUSIONS: Intraluminally or topically delivered statins reduced IH. The efficacy of single-dose, locally delivered statin alone may lead to novel treatments to prevent IH. The different routes of administration may allow for treatment during endovascular procedures, without the need for systemic therapy.

The role of assembly parameters on polyplex poly(beta-amino ester) nanoparticle transfections.

Intracellular delivery of nucleic acids to mammalian cells using polyplex nanoparticles (NPs) remains a challenge both in vitro and in vivo, with transfections often suffering from variable efficacy. To improve reproducibility and efficacy of transfections in vitro using a next-generation polyplex transfection material poly(beta-amino ester)s (PBAEs), the influence of multiple variables in the preparation of these NPs on their transfection efficacy was explored. The results indicate that even though PBAE/pDNA polyplex NPs are formed by the self-assembly of polyelectrolytes, their transfection is not affected by the manner in which the components are mixed, facilitating self-assembly in a single step, but timing for self-assembly of 5-20 min is optimal. In addition, even though the biomaterials are biodegradable in water, their efficacy is not affected by up to eight freeze-thaw cycles of the polymer. It was found that there is a greater stability of nucleic acid-complexed polymer as a polyplex nanoparticle compared with free polymer. Finally, by exploring multiple buffer systems, it was identified that utilization of divalent cation magnesium or calcium acetate buffers at pH 5.0 is optimal for transfection using these polymeric materials, boosting transfection several folds compared with monovalent cations. Together, these results can improve the reproducibility and efficacy of PBAE and similar polyplex nanoparticle transfections and improve the robustness of using these biomaterials for bioengineering and biotechnology applications.

Knee osteoarthritis patients with more subchondral cysts have altered tibial subchondral bone mineral density.

BACKGROUND: Subchondral bone cysts are a widely observed, but poorly understood, feature in patients with knee osteoarthritis (OA). Clinical quantitative computed tomography (QCT) has the potential to characterize cysts in vivo but it is unclear which specific cyst parameters (e.g., number, size) are associated with clinical signs of OA, such as disease severity or pain. The objective of this study was to use QCT-based image-processing techniques to characterize subchondral tibial cysts in patients with knee OA and to explore relationships between proximal tibial subchondral cyst parameters and subchondral bone density as well as clinical characteristics of OA (alignment, joint space narrowing (JSN), OA severity, pain) in patients with knee OA. METHODS: The preoperative knee of 42 knee arthroplasty patients was scanned using QCT. Patient characteristics were obtained, including OA severity, knee pain, JSN, and alignment. We used 3D image processing techniques to obtain cyst parameters including: cyst number, cyst number per proximal tibial volume, cyst volume per proximal tibial volume, as well as maximum and average cyst volume across the proximal tibia, as well as regional bone mineral density (BMD) excluding cysts. We used Spearman's correlation coefficients to explore associations between patient characteristics and cyst parameters. RESULTS: At both the medial and lateral compartments of the proximal tibia, greater cyst number and volume were associated with higher BMD. At the lateral region, cyst number and volume were also associated with lateral OA severity, lateral JSN, alignment and sex. Pain was not associated with any cyst parameters at any region. CONCLUSION: Cyst number and volume were associated with BMD at both the medial and lateral compartments. Lateral cyst number and volume were also associated with joint alignment, OA severity, JSN and sex. This is the first study to use clinical QCT to explore subchondral tibial cysts in patients with knee OA and provides further evidence of the relationships between subchondral cysts and clinical OA characteristics.

Bioreducible Polymeric Nanoparticles Containing Multiplexed Cancer Stem Cell Regulating miRNAs Inhibit Glioblastoma Growth and Prolong Survival.

Despite our growing molecular-level understanding of glioblastoma (GBM), treatment modalities remain limited. Recent developments in the mechanisms of cell fate regulation and nanomedicine provide new avenues by which to treat and manage brain tumors via the delivery of molecular therapeutics. Here, we have developed bioreducible poly(ß-amino ester) nanoparticles that demonstrate high intracellular delivery efficacy, low cytotoxicity, escape from endosomes, and promotion of cytosol-targeted environmentally triggered cargo release for miRNA delivery to tumor-propagating human cancer stem cells. In this report, we combined this nanobiotechnology with newly discovered cancer stem cell inhibiting miRNAs to develop self-assembled miRNA-containing polymeric nanoparticles (nano-miRs) to treat gliomas. We show that these nano-miRs effectively intracellularly deliver single and combination miRNA mimics that inhibit the stem cell phenotype of human GBM cells in vitro. Following direct intratumoral infusion, these nano-miRs were found to distribute through the tumors, inhibit the growth of established orthotopic human GBM xenografts, and cooperatively enhance the response to standard-of-care γ radiation. Co-delivery of two miRNAs, miR-148a and miR-296-5p, within the bioreducible nano-miR particles enabled long-term survival from GBM in mice.

Contrast-enhanced computed tomography (CECT) attenuation is associated with stiffness of intact knee cartilage.

Contrast-enhanced computed tomography (CECT) using charged contrast-agents enables quantification of cartilage glycosaminoglycan content. Since glycosaminoglycan content is a key determinant of cartilage compressive stiffness, CECT measurements have the potential to non-invasively assess cartilage stiffness. The objective of this study was to determine whether CECT attenuation, using a cationic contrast-agent (CA4+), correlates with the stiffness of intact cartilage. Six fresh femoral and six fresh tibial compartments with intact cartilage were obtained from patients undergoing total knee replacement surgery. The instantaneous stiffness was determined for 25-50 points on the surface of each compartment using an established indentation technique. The samples were then immersed in CA4+ solution for 48 h, scanned in a micro-CT scanner, and the average CECT attenuation at each indentation site was found for the superficial cartilage. A significant (p < 0.01) and positive correlation was observed between stiffness and CECT attenuation for sites from each individual cartilage surface, with correlation coefficients ranging from r = 0.37-0.57 and r = 0.48-0.69 (p < 0.01) for the tibia and femur, respectively. When data for each type of cartilage surface were pooled together, the correlation coefficients were r = 0.73 for femoral condyle data points and r = 0.49 for tibial plateau data points. CECT provided a map of cartilage stiffness across each surface, which allows regions of low stiffness to be identified. These findings support continued evaluation and development of quantitative imaging techniques to assess the functional properties of cartilage. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2641-2647, 2018.

Biodegradable STING agonist nanoparticles for enhanced cancer immunotherapy.

Therapeutic cancer vaccines require adjuvants leading to robust type I interferon and proinflammatory cytokine responses in the tumor microenvironment to induce an anti-tumor response. Cyclic dinucleotides (CDNs), a potent Stimulator of Interferon Receptor (STING) agonist, are currently in phase I trials. However, their efficacy may be limited to micromolar concentrations due to the cytosolic residence of STING in the ER membrane. Here we utilized biodegradable, poly(beta-amino ester) (PBAE) nanoparticles to deliver CDNs to the cytosol leading to robust immune response at >100-fold lower extracellular CDN concentrations in vitro. The leading CDN PBAE nanoparticle formulation induced a log-fold improvement in potency in treating established B16 melanoma tumors in vivo when combined with PD-1 blocking antibody in comparison to free CDN without nanoparticles. This nanoparticle-mediated cytosolic delivery method for STING agonists synergizes with checkpoint inhibitors and has strong potential for enhanced cancer immunotherapy.

Surface engineering for lymphocyte programming.

The once nascent field of immunoengineering has recently blossomed to include approaches to deliver and present biomolecules to program diverse populations of lymphocytes to fight disease. Building upon improved understanding of the molecular and physical mechanics of lymphocyte activation, varied strategies for engineering surfaces to activate and deactivate T-Cells, B-Cells and natural killer cells are in preclinical and clinical development. Surfaces have been engineered at the molecular level in terms of the presence of specific biological factors, their arrangement on a surface, and their diffusivity to elicit specific lymphocyte fates. In addition, the physical and mechanical characteristics of the surface including shape, anisotropy, and rigidity of particles for lymphocyte activation have been fine-tuned. Utilizing these strategies, acellular systems have been engineered for the expansion of T-Cells and natural killer cells to clinically relevant levels for cancer therapies as well as engineered to program B-Cells to better combat infectious diseases.

A Triple-Fluorophore-Labeled Nucleic Acid pH Nanosensor to Investigate Non-viral Gene Delivery.

There is a need for new tools to better quantify intracellular delivery barriers in high-throughput and high-content ways. Here, we synthesized a triple-fluorophore-labeled nucleic acid pH nanosensor for measuring intracellular pH of exogenous DNA at specific time points in a high-throughput manner by flow cytometry following non-viral transfection. By including two pH-sensitive fluorophores and one pH-insensitive fluorophore in the nanosensor, detection of pH was possible over the full physiological range. We further assessed possible correlation between intracellular pH of delivered DNA, cellular uptake of DNA, and DNA reporter gene expression at 24 hr post-transfection for poly-L-lysine and branched polyethylenimine polyplex nanoparticles. While successful transfection was shown to clearly depend on median cellular pH of delivered DNA at the cell population level, surprisingly, on an individual cell basis, there was no significant correlation between intracellular pH and transfection efficacy. To our knowledge, this is the first reported instance of high-throughput single-cell analysis between cellular uptake of DNA, intracellular pH of delivered DNA, and gene expression of the delivered DNA. Using the nanosensor, we demonstrate that the ability of polymeric nanoparticles to avoid an acidic environment is necessary, but not sufficient, for successful transfection.