Reciprocal regulation of pancreatic ductal adenocarcinoma growth and molecular subtype by HNF4α and SIX1/4.

OBJECTIVE: Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with a 5-year survival of less than 5%. Transcriptomic analysis has identified two clinically relevant molecular subtypes of PDAC: classical and basal-like. The classical subtype is characterised by a more favourable prognosis and better response to chemotherapy than the basal-like subtype. The classical subtype also expresses higher levels of lineage specifiers that regulate endodermal differentiation, including the nuclear receptor hepatocyte nuclear factor 4 α (HNF4α). The objective of this study is to evaluate the role of HNF4α, SIX4 and SIX1 in regulating the growth and molecular subtype of PDAC. DESIGN: We manipulate the expression of HNF4α, SIX4 and SIX1 in multiple in vitro and in vivo PDAC models. We determine the consequences of manipulating these genes on PDAC growth, differentiation and molecular subtype using functional assays, gene expression analysis and cross-species comparisons with human datasets. RESULTS: We show that HNF4α restrains tumour growth and drives tumour cells toward an epithelial identity. Gene expression analysis of murine models and human tumours shows that HNF4α activates expression of genes associated with the classical subtype. HNF4α also directly represses SIX4 and SIX1, two mesodermal/neuronal lineage specifiers expressed in the basal-like subtype. Finally, SIX4 and SIX1 drive proliferation and regulate differentiation in HNF4α-negative PDAC. CONCLUSION: Our data show that HNF4α regulates the growth and molecular subtype of PDAC by multiple mechanisms, including activation of the classical gene expression programme and repression of SIX4 and SIX1, which may represent novel dependencies of the basal-like subtype.

Drug-delivering nerve conduit improves regeneration in a critical-sized gap.

Autologous nerve grafts are the current "gold standard" for repairing large nerve gaps. However, they cause morbidity at the donor nerve site and only a limited amount of nerve can be harvested. Nerve conduits are a promising alternative to autografts and can act as guidance cues for the regenerating axons, without the need to harvest donor nerve. Separately, it has been shown that localized delivery of GDNF can enhance axon growth and motor recovery. FK506, an FDA approved small molecule, has also been shown to enhance peripheral nerve regeneration. This paper describes the design of a novel hole-based drug delivery apparatus integrated with a polytetrafluoroethylene (PTFE) nerve conduit for controlled local delivery of a protein such as GDNF or a small molecule such as FK506. The PTFE devices were tested in a diffusion chamber, and the bioactivity of the released media was evaluated by measuring neurite growth of dorsal root ganglions (DRGs) exposed to the released drugs. The drug delivering nerve guide was able to release bioactive concentrations of FK506 or GDNF. Following these tests, optimized drug releasing nerve conduits were implanted across 10 mm sciatic nerve gaps in a BL6 yellow fluorescent protein (YFP) mouse model, where they demonstrated significant improvement in muscle mass, compound muscle action potential, and axon myelination in vivo as compared with nerve conduits without the drug. The drug delivery nerve guide could release drug for extended periods of time and enhance axon growth in vitro and in vivo.

Local FK506 delivery at the direct nerve repair site improves nerve regeneration.

INTRODUCTION: The objective of this study is to assess the efficacy of local tacrolimus (FK506) delivery to improve outcomes in the setting of nerve transection injury. METHODS: FK506 embedded poly(lactide-co-caprolactone) films capable of extended, localized release of FK506 were developed. FK506 rate of release testing and bioactivity assay was performed. Mouse sciatic nerve transection and direct repair model was used to evaluate the effect extended, local delivery of FK506 had on nerve regeneration outcomes. RESULTS: Linear release of FK506 was observed for 30 days and released FK506 matched control levels of neurite extension in the dorsal root ganglion assay. Groups treated with local FK506 had greater gastrocnemius muscle weight, foot electromyogram, and number of axons distal of the repair site than non-FK506 groups. DISCUSSION: Results of this study indicate that extended, localized delivery of FK506 to nerve injuries can improve nerve regeneration outcomes in a mouse sciatic nerve transection and repair.

Glitazone loaded fat enhances adiponectin production and inhibits breast cancer cell proliferation.

Obesity and diabetes are both associated risk factors for developing breast cancer and poor patient outcomes. Adipose cells are an important endocrine system and are the main producer of adiponectin, with lean patients having higher circulating levels. Patients with diabetes are often treated with thiazolidinediones, glitazones, which also increase adiponectin production. Importantly high circulating levels of adiponectin and treatment with glitazone are associated with increased breast cancer patient survival. This study investigates the potential of using adipose tissue laden with glitazones to act as a drug depot, increase adiponectin levels, and locally release glitazones to inhibit breast cancer cell proliferation. The breast cancer cell lines MCF-7 and MBA-MD-231, and the normal breast epithelial cell line MCF-10A were exposed to media containing a range of concentrations of recombinant adiponectin, pioglitazone, or conditioned media obtained from pioglitazone laden adipose tissue to determine the impact of the different treatments on cell proliferation. The MCF-7 cells demonstrated the greatest reduction in proliferation upon exposure to adiponectin and pioglitazone with lower reductions observed in the MDA-MD-231 and MCF-10a cell lines. All three cell lines exhibited reductions in proliferation in the presence of pioglitazone loaded adipose tissue. Additionally, adiponectin and pioglitazone levels were higher in the media from glitazone loaded adipose tissue. Drug loaded adipose tissue could potentially be used to deliver adiponectin and glitazone to breast cancer cells and inhibit proliferation. Future research will examine the potential efficacy of this treatment approach in vivo.

Effect Of combining FK506 and neurotrophins on neurite branching and elongation.

INTRODUCTION: There is a clinical need to improve the outcomes of peripheral nerve regeneration and repair after injury. In addition to its immunosuppressive effects, FK506 (tacrolimus) has been shown to have neuroregenerative properties. To determine biologically relevant local FK506 and growth factor concentrations, we performed an in vitro bioassay using dorsal root ganglion (DRG) from chicken embryos. METHODS: Neurite elongation and neurite branching were analyzed microscopically after addition of FK506, glial cell line-derived neurotrophic factor (GDNF), and nerve growth factor (NGF), each alone and in combination. RESULTS: FK506 induced modest neurite elongation (∼500-800 µm) without improving neurite branching significantly. The combination of FK506 with NGF, GDNF, or both, exerted a potentiating or competitive effect on neurite elongation (∼700-1100 µm) based on dosage and competitive effect on neurite branching (∼0.2-0.4). CONCLUSIONS: These results strongly suggest that the interaction of FK506 with GDNF and NGF mediates distinct enhancement of neurite growth. Muscle Nerve 55: 570-581, 2017.

Lineage tracing reveals the dynamic contribution of Hes1+ cells to the developing and adult pancreas.

Notch signaling regulates numerous developmental processes, often acting either to promote one cell fate over another or else to inhibit differentiation altogether. In the embryonic pancreas, Notch and its target gene Hes1 are thought to inhibit endocrine and exocrine specification. Although differentiated cells appear to downregulate Hes1, it is unknown whether Hes1 expression marks multipotent progenitors, or else lineage-restricted precursors. Moreover, although rare cells of the adult pancreas express Hes1, it is unknown whether these represent a specialized progenitor-like population. To address these issues, we developed a mouse Hes1(CreERT2) knock-in allele to inducibly mark Hes1(+) cells and their descendants. We find that Hes1 expression in the early embryonic pancreas identifies multipotent, Notch-responsive progenitors, differentiation of which is blocked by activated Notch. In later embryogenesis, Hes1 marks exocrine-restricted progenitors, in which activated Notch promotes ductal differentiation. In the adult pancreas, Hes1 expression persists in rare differentiated cells, particularly terminal duct or centroacinar cells. Although we find that Hes1(+) cells in the resting or injured pancreas do not behave as adult stem cells for insulin-producing beta (ß)-cells, Hes1 expression does identify stem cells throughout the small and large intestine. Together, these studies clarify the roles of Notch and Hes1 in the developing and adult pancreas, and open new avenues to study Notch signaling in this and other tissues.

Side-to-side nerve bridges reduce muscle atrophy after peripheral nerve injury in a rodent model.

BACKGROUND: Peripheral nerve injury can result in muscle atrophy and long-term disability. We hypothesize that creating a side-to-side bridge to link an injured nerve with a healthy nerve will reduce muscle atrophy and improve muscle function. METHODS: Sprague-Dawley rats were divided into four groups (n = 7 per group). Group 1: transection only--a 10-mm gap was created in the proximal tibial nerve; group 2: transected plus repaired--the transected tibial nerve was repaired; group 3: transected plus repaired plus nerve bridge--transected nerve repaired with a distal nerve bridge between the tibial and peroneal nerves via epineurial windows; and group 4: transected plus nerve bridge--transected tibial nerve left unrepaired and distal bridge added. Gait was assessed every 2 wk. At 90 d the following measures were determined: gastrocnemius mass, muscle and nerve nuclear density, and axonal infiltration into the nerve bridge. RESULTS: Groups 3 and 4 had greater improvements in walking track recovery than groups 1 and 2. Group 3's gastrocnemius muscles exhibited the least amount of atrophy. Groups 1, 2, and 4 exhibited greater histologic appearance of muscle breakdown compared with group 3 and control muscle. Finally, most bridges in groups 3 and 4 had neuronal sprouting via the epineurial windows. CONCLUSIONS: Our study demonstrated reduced muscle atrophy with a side-to-side nerve bridge in the setting of peripheral nerve injury. These results support the application of novel side-to-side bridges in combination with traditional end-to-end neurorrhaphy to preserve muscle viability after peripheral nerve injuries.

Sintered fluorapatite scaffolds as an autograft-like engineered bone graft.

Hydroxyapatite (HA)-based materials are widely used as bone substitutes due to their inherent biocompatibility, osteoconductivity, and bio-absorption properties. However, HA scaffolds lack compressive strength when compared to autograft bone. It has been shown that the fluoridated form of HA, fluorapatite (FA), can be sintered to obtain this desired strength as well as slower degradation properties. Also, FA surfaces have been previously shown to promote stem cell differentiation toward an osteogenic lineage. Thus, it was hypothesized that FA, with and without stromal vascular fraction (SVF), would guide bone healing to an equal or better extent than the clinical gold standard. The regenerative potentials of these scaffolds were tested in 32 Lewis rats in a femoral condylar defect model with untreated (negative), isograft (positive), and commercial HA as controls. Animals were survived for 12 weeks post-implantation. A semi-quantitative micro-CT analysis was developed to quantify the percent new bone formation within the defects. Our model showed significantly higher (p < .05) new bone depositions in all apatite groups compared to the autograft group. Overall, the FA group had the most significant new bone deposition, while the differences between HA, FA, and FA + SVF were insignificant (p > .05). Histological observations supported the micro-CT findings and highlighted the presence of healthy bone tissues without interposing capsules or intense immune responses for FA groups. Most importantly, the regenerating bone tissue within the FA + SVF scaffolds resembled the architecture of the surrounding trabecular bone, showing intertrabecular spaces, while the FA group presented a denser cortical bone-like architecture. Also, a lower density of cells was observed near FA granules compared to HA surfaces, suggesting a reduced immune response. This first in vivo rat study supported the tested hypothesis, illustrating the utility of FA as a bone scaffold material.

A Novel Vascular Anastomotic Coupling Device for End-to-End Anastomosis of Arteries and Veins.

OBJECTIVE: Hand-sutured (HS) techniques remain the gold standard for most microvascular anastomoses in microsurgery. HS techniques can result in endothelial lacerations and back wall suturing, leading to complications such as thrombosis and free tissue loss. A novel force-interference-fit vascular coupling device (FIF-VCD) system can potentially reduce the need for HS and improve end-to-end anastomosis. This study aims to describe the development and testing of a novel FIF-VCD system for 1.5 to 4.0 mm outside diameter arteries and veins. METHODS: Benchtop anastomoses were performed using porcine cadaver arteries and veins. Decoupling force and anastomotic leakage were tested under simulated worst-case intravital physiological conditions. The 1.5 mm FIF-VCD system was used to perform cadaver rat abdominal aorta anastomoses. RESULTS: Benchtop testing showed that the vessels coupled with the FIF-VCD system could withstand simulated worst-case intravital physiological conditions with a 95% confidence interval for the average decoupling force safety factor of 8.2 ± 1.0 (5.2 ± 1.0 N) and a 95% confidence interval for the average leakage rate safety factor of 26 ± 3.6 (8.4 ± 0.14 and 95 ± 1.4 µL/s at 150 and 360 mmHg, respectively) when compared to HS anastomotic leakage rates (310 ± 14 and 2,100 ± 72 µL/s at 150 and 360 mmHg, respectively). The FIF-VCD system was successful in performing cadaver rat abdominal aorta anastomoses. CONCLUSION: The FIF-VCD system can potentially replace HS in microsurgery, allowing the safe and effective connection of arteries and veins. Further studies are needed to confirm the clinical viability and effectiveness of the FIF-VCD system.

A Non-Contrast Multi-Parametric MRI Biomarker for Assessment of MR-Guided Focused Ultrasound Thermal Therapies.

OBJECTIVE: We present the development of a non-contrast multi-parametric magnetic resonance (MPMR) imaging biomarker to assess treatment outcomes for magnetic resonance-guided focused ultrasound (MRgFUS) ablations of localized tumors. Images obtained immediately following MRgFUS ablation were inputs for voxel-wise supervised learning classifiers, trained using registered histology as a label for thermal necrosis. METHODS: VX2 tumors in New Zealand white rabbits quadriceps were thermally ablated using an MRgFUS system under 3 T MRI guidance. Animals were re-imaged three days post-ablation and euthanized. Histological necrosis labels were created by 3D registration between MR images and digitized H&E segmentations of thermal necrosis to enable voxel-wise classification of necrosis. Supervised MPMR classifier inputs included maximum temperature rise, cumulative thermal dose (CTD), post-FUS differences in T2-weighted images, and apparent diffusion coefficient, or ADC, maps. A logistic regression, support vector machine, and random forest classifier were trained in red a leave-one-out strategy in test data from four subjects. RESULTS: In the validation dataset, the MPMR classifiers achieved higher recall and Dice than a clinically adopted 240 cumulative equivalent minutes at 43 °C (CEM 43) threshold (0.43) in all subjects. The average Dice scores of overlap with the registered histological label for the logistic regression (0.63) and support vector machine (0.63) MPMR classifiers were within 6% of the acute contrast-enhanced non-perfused volume (0.67). CONCLUSIONS: Voxel-wise registration of MPMR data to histological outcomes facilitated supervised learning of an accurate non-contrast MR biomarker for MRgFUS ablations in a rabbit VX2 tumor model.

Efficacy of sintered Zinc-doped fluorapatite scaffold as an antimicrobial regenerative bone filler for dental applications.

OBJECTIVES: The objective of this study was to assess whether zinc-doped fluorapatite (ZnFA) could serve as an effective antimicrobial dental bone filler for bone regeneration compared to autografts. METHODS: FA and 2 % zinc-doped FA (2ZnFA) were synthesized and characterized in-house. Compressed and sintered FA and 2ZnFA disks were incubated with bacteria to assess antimicrobial properties. Adipose-derived stem cells were cultured on these discs to evaluate the surfaces' ability to support cell growth and promote osteogenic differentiation. Surfaces exhibiting the highest expressions of the bone markers osteopontin and osteocalcin were selected for an in vivo study in a rat mandibular defect model. Twenty rats were divided into 5 groups, equally, and a 5 mm surgical defect of the jaw was left untreated or filled with 2ZnFA, FA, autograft, or demineralized bone matrix (DBM). At 12 weeks, the defects and surrounding tissues were harvested and subjected to microCT and histological evaluations. RESULTS: Standard techniques such as FTIR, ICP-MS, fluoride probe, and XRD revealed the sintered FA and ZnFA's chemical compositions and structures. Bacterial studies revealed no significant differences in surface bacterial adhesion properties between FA and 2ZnFA, but significantly fewer bacterial loads than control titanium discs (p < 0.05). Cell culture data confirmed that both surfaces could support cell growth and promote the osteogenic differentiation of stem cells. MicroCT analysis confirmed statistical similarities in bone regeneration within FA, 2ZnFA, and autograft groups. CONCLUSION: The data suggests that both FA and 2ZnFA could serve as alternatives to autograft materials, which are the current gold standard. Moreover, these bone fillers outperformed DBM, an allograft material commonly used as a dental bone void filler. CLINICAL SIGNIFICANCE: The use of FA or 2ZnFA for treating mandibular defects led to bone regeneration statistically similar to autograft repair and significantly outperformed the widely used dental bone filler, DBM. Additional translational research may confirm FA-based materials as superior substitutes for existing synthetic bone fillers, ultimately enhancing patient outcomes.

A ropivacaine-eluting poly(lactide-co-caprolactone) wound dressing provided enhanced analgesia in partial-thickness porcine injuries.

BACKGROUND: Partial-thickness skin wounds are some of the most painful injuries due to large areas of exposed nerve endings. These injuries often require systemic opioid treatments to manage pain adequately. However, in 2021 alone, the CDC reported nearly 17,000 prescription opioid-related deaths in the USA, highlighting the ongoing need for non-opioid treatment strategies. In this manuscript, we developed a novel single-application ropivacaine-eluting primary wound dressing that could provide sustained ropivacaine delivery to partial-thickness wounds and assessed its in vivo feasibility for prolonged non-opioid analgesia. METHODS: Sustained release of ropivacaine from a poly(lactide-co-e-caprolactone) matrix was first optimized in vitro using dissolution testing and a Box Behnken design of experiments. The optimized dressing was then tested against a clinical control silicone dressing in a porcine partial-thickness wound study to assess analgesic effect, pharmacokinetics, and wound healing. RESULTS: The ropivacaine-eluting dressing showed a moderate analgesic effect in vivo, where normalized single pinprick scores significantly improved pain over the testing period (4-168h) (control vs treatment: 232±25% vs 145±16%, p<0.0003). Ropivacaine blood plasma levels peaked at 8 hours post-treatment, with a maximum concentration of 246 ± 74 ng/mL. No significant differences in wound healing were found when compared to control. CONCLUSION: The ropivacaine-loaded poly(lactide-co-e-caprolactone)-based wound dressing provided sustained delivery of ropivacaine to partial-thickness skin wounds and enhanced analgesic effect compared to a clinical standard control dressing.

Local delivery of FK506 to a nerve allograft is comparable to systemic delivery at suppressing allogeneic graft rejection.

The objective of this study was to determine if locally delivered FK506 could prevent allogeneic nerve graft rejection long enough to allow axon regeneration to pass through the nerve graft. An 8mm mouse sciatic nerve gap injury repaired with a nerve allograft was used to assess the effectiveness of local FK506 immunosuppressive therapy. FK506-loaded poly(lactide-co-caprolactone) nerve conduits were used to provide sustained local FK506 delivery to nerve allografts. Continuous and temporary systemic FK506 therapy to nerve allografts, and autograft repair were used as control groups. Serial assessment of inflammatory cell and CD4+ cell infiltration into the nerve graft tissue was performed to characterize the immune response over time. Nerve regeneration and functional recovery was serially assessed by nerve histomorphometry, gastrocnemius muscle mass recovery, and the ladder rung skilled locomotion assay. At the end of the study, week 16, all the groups had similar levels of inflammatory cell infiltration. The local FK506 and continuous systemic FK506 groups had similar levels of CD4+ cell infiltration, however, it was significantly greater than the autograft control. In terms of nerve histmorphometry, the local FK506 and continunous systemic FK506 groups had similar amounts of myelinated axons, although they were significantly lower than the autograft and temporary systemic FK506 group. The autograft had significantly greater muscle mass recovery than all the other groups. In the ladder rung assay, the autograft, local FK506, and continuous systemic FK506 had similar levels of skilled locomotion performance, whereas the temporary systemic FK506 group had significanty better performance than all the other groups. The results of this study suggest that local delivery of FK506 can provide comparable immunosuppression and nerve regeneration outcomes as systemically delivered FK506.

Fluorapatite and fluorohydroxyapatite apatite surfaces drive adipose-derived stem cells to an osteogenic lineage.

PURPOSE: Hydroxyapatite (HA) scaffolds are common replacement materials used in the clinical management of critical-sized bone defects. This study was undertaken to examine the potential benefits of fluoridated derivatives of hydroxyapatite, fluorapatite (FA), and fluorohydroxyapatite (FHA) as bone scaffolds in conjunction with adipose-derived stem cells (ADSCs). If FHA and FA surfaces could drive the differentiation of stem cells to an osteogenic phenotype, the combination of these ceramic scaffolds with ADSCs could produce materials with mechanical strength and remodeling potential comparable to autologous bone. This study was designed to investigate the ability of the apatite surfaces HA, FA, and FHA produced at different sintering temperatures to drive ADSCs toward osteogenic lineages. METHODS: HA, FHA, and FA surfaces sintered at 1150 °C and 1250 °C were seeded with ADSCs and evaluated for cell growth and gene and protein expression of osteogenic markers at 2 and 10 days post-seeding. RESULTS: In vitro, ADSC cells were viable on all surfaces; however, differentiation of these cells into osteoblastic lineage only observed in apatite surfaces. ADSCs seeded on FA and FHA expressed genes and proteins related to osteogenic differentiation markers to a greater extent by Day 2 when compared to HA and cell culture controls. By day 10, HA, FA, and FHA all expressed more bone differentiation markers compared to cell culture controls. CONCLUSION: FA and FHA apatite scaffolds may promote the differentiation of ADSCs at an earlier time point than HA surfaces. Combining apatite scaffolds with ADSCs has the potential to improve bone regeneration following bone injury.

Evaluating the influence of particle morphology and density on the viscosity and injectability of a novel long-acting local anesthetic suspension.

Proper pain management is well understood to be one of the fundamental aspects of a healthy postoperative recovery in conjunction with mobility and nutrition. Approximately, 10% of patients prescribed opioids after surgery continue to use opioids in the long-term and as little as 10 days on opioids can result in addiction. In an effort to provide physicians with an alternative pain management technique, this work evaluates the material properties of a novel local anesthetic delivery system designed for controlled release of bupivacaine for 72 hours. The formulation utilizes solid-lipid microparticles that encapsulate the hydrophobic molecule bupivacaine in its free-base form. The lipid microparticles are suspended in a non-crosslinked hyaluronic acid hydrogel, which acts as the microparticle carrier. Two different particle manufacturing techniques, milling and hot homogenization, were evaluated in this work. The hot homogenized particles had a slower and more controlled release than the milled particles. Rheological techniques revealed that the suspension remains a viscoelastic fluid when loaded with either particle type up to 25% (w/v) particles densities. Furthermore, the shear thinning properties of the suspension media, hyaluronic acid hydrogel, were conserved when bupivacaine-loaded solid-lipid microparticles were loaded up to densities of 25% (w/v) particle loading. The force during injection was measured for suspension formulations with varying hyaluronic acid hydrogel concentrations, particle densities, particle types and particle sizes. The results indicate that the formulation viscosity is highly dependent on particle density, but hyaluronic acid hydrogel is required for lowering injection forces as well as minimizing clogging events.

Genexol inhibits primary tumour growth and metastases in gemcitabine-resistant pancreatic ductal adenocarcinoma.

INTRODUCTION: Gemcitabine, the current standard of care for pancreatic ductal adenocarcinoma (PDA), has a less than 10% partial response rate. Genexol-PM, a modified form of paclitaxel, has been shown to have antitumour effects in clinical trials of metastatic breast and small-lung-cell carcinoma. The aim of the present study was to determine if Genexol would be a beneficial treatment for gemcitabine-resistant PDA. MATERIALS AND METHODS: We measured the in vitro IC50s of gemcitabine and genexol in cell lines sensitive and resistant to gemcitabine. In vivo, animals with orthotopic pancreatic tumours, resistant to gemcitabine, were treated with phosphate-buffered saline (PBS), gemcitabine, Genexol or gemcitabine+Genexol. Tumour progression was monitored using red fluorescent protein imaging. RESULTS: We showed equivalent IC50s for gemcitabine-sensitive and gemcitabine-resistant cell lines when treated with genexol. In vivo treatment with genexol resulted in a greater per cent reduction in tumour size, less metastatic spread and longer survival compared with treatment with gemcitabine. DISCUSSION: Genexol proved to be an effective treatment for gemcitabine-resistant PDA. These data combined with the successful clinical use of genexol in Phase II trials of other malignancies suggests it maybe an effective treatment for pancreatic cancer, specifically for those patients resistant to gemcitabine.

Compression of the vascular wall to create a friction fit in a vascular anastomotic coupler.

A previously reported microvascular coupler was shown to effectively create vascular anastomoses, but was too large for practical clinical use. To safely reduce coupler size, certain failure modes needed to be better understood. The coupler functions, in part, by compressing the vessel wall between two concentric rings, creating a friction fit that anchors the device to the vessel. This work investigates the relationship between vessel wall compression and resulting friction fit strength to ensure reducing coupler size will not unduly increase the risk that this friction fit might fail. Vascular walls were compressed to a specified strain and the tensile force required to overcome the resulting friction was measured. Experiments were conducted with various vessel types (Porcine common carotid artery, splenic artery, and jugular vein), across a range of compressive strains (55-95%), and by using either PEEK or HDPE to compress the vessel. Tensile force was increased at a rate of 5 g/min or held constant for 24 h. For experiments with incrementally increasing force, the force at failure varied with compressive strain via a power function. At 70% compression, PEEK produced 4.6 times stronger friction fits than HDPE, and common carotid arteries and splenic arteries produced 1.8 and 1.3 times stronger fits than jugular veins respectively. For experiments where tensile force was applied for 24 h, much lower forces were required to overcome friction. These results were compared to friction fit failure in a coupler prototype and it was found that the prototypes failed at just 30% of the force required to cause vessel slip under the other test conditions. These results were used to develop a model that predicts the probability of device failure via vessel slipping (one design, smaller than previously reported, was estimated to fail at maximum in vivo axial stress once in 500 anastomoses, a potentially safe level of risk).

Modeling diffusion-based drug release inside a nerve conduit in vitro and in vivo validation study.

The objective of this work was to develop a model and understand the diffusion of a drug into and throughout a drug delivering nerve conduit from a surrounding reservoir through a hole in the wall separating the lumen of the conduit and the reservoir. A mathematical model based on Fick's law of diffusion was developed using the finite difference method to understand the drug diffusion and the effect of varying device parameters on the concentration of drug delivered from a hole-based drug delivery device. The mathematical model was verified using a physical microfluidic (µFD) model and an in vitro/in vivo release test using prototype devices. The results of the mathematical model evaluation and microfluidic device testing offered positive insight into the reliability and function of the reservoir and hole-based drug delivering nerve conduit. The mathematical model demonstrated how changing device parameters would change the drug concentration inside the device. It was observed that the drug release in the conduit could be tuned by both concentration scaling and changing the hole size or number of holes. Based on the results obtained from the microfluidic device, the error in the mathematical drug release model was shown to be less than 10% when comparing the data obtained from mathematical model and µFD model. The data highlights the flexibility of having a hole-based drug delivery system, since the drug release can be scaled predictably by changing the device parameters or the concentration of the drug in the reservoir. Graphical abstract .

Learning Multiparametric Biomarkers for Assessing MR-Guided Focused Ultrasound Treatment of Malignant Tumors.

Noninvasive MR-guided focused ultrasound (MRgFUS) treatments are promising alternatives to the surgical removal of malignant tumors. A significant challenge is assessing the viability of treated tissue during and immediately after MRgFUS procedures. Current clinical assessment uses the nonperfused volume (NPV) biomarker immediately after treatment from contrast-enhanced MRI. The NPV has variable accuracy, and the use of contrast agent prevents continuing MRgFUS treatment if tumor coverage is inadequate. This work presents a novel, noncontrast, learned multiparametric MR biomarker that can be used during treatment for intratreatment assessment, validated in a VX2 rabbit tumor model. A deep convolutional neural network was trained on noncontrast multiparametric MR images using the NPV biomarker from follow-up MR imaging (3-5 days after MRgFUS treatment) as the accurate label of nonviable tissue. A novel volume-conserving registration algorithm yielded a voxel-wise correlation between treatment and follow-up NPV, providing a rigorous validation of the biomarker. The learned noncontrast multiparametric MR biomarker predicted the follow-up NPV with an average DICE coefficient of 0.71, substantially outperforming the current clinical standard (DICE coefficient = 0.53). Noncontrast multiparametric MR imaging integrated with a deep convolutional neural network provides a more accurate prediction of MRgFUS treatment outcome than current contrast-based techniques.