Evaluation of permeability and fluid wicking in woven fiber bone scaffolds.

Research characterizing transport of nutrients and waste in tissue engineering scaffolds has led to the study of scaffold properties that contribute to permeability and porosity of the scaffold. Both permeability and porosity contribute to the transport properties of the scaffold; however, permeability relates to the degree to which pores are interconnected within the scaffold. This work evaluated permeability for woven polymer fiber scaffolds by modulating the following scaffold parameters: material combination, weave configuration, and fiber geometry. Materials tested were poly-l-lactide and poly-l-lactide-co-ɛ-caprolactone in various combinations. Plain and crowfoot weave configurations were compared, and grooved wicking fibers were compared with round cross-section fibers to study fiber geometry. A modification of the constant head hydraulic conductivity test was used in combination with a vertical wicking test to determine levels of permeability of the woven scaffolds. Results showed a significant effect on permeability for combinations of weave configuration, fiber geometry, and material combination. However, modulating fiber geometry demonstrated the most significant contribution to permeability. This result suggests the grooved wicking geometry may be used in scaffold development to regulate transport by selectively moving fluid away or toward the area of interest by capillary action. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 107B: 306-313, 2019.

Characterization and Separation of Cancer Cells with a Wicking Fiber Device.

Current cancer diagnostic methods lack the ability to quickly, simply, efficiently, and inexpensively screen cancer cells from a mixed population of cancer and normal cells. Methods based on biomarkers are unreliable due to complexity of cancer cells, plasticity of markers, and lack of common tumorigenic markers. Diagnostics are time intensive, require multiple tests, and provide limited information. In this study, we developed a novel wicking fiber device that separates cancer and normal cell types. To the best of our knowledge, no previous work has used vertical wicking of cells through fibers to identify and isolate cancer cells. The device separated mouse mammary tumor cells from a cellular mixture containing normal mouse mammary cells. Further investigation showed the device separated and isolated human cancer cells from a heterogeneous mixture of normal and cancerous human cells. We report a simple, inexpensive, and rapid technique that has potential to identify and isolate cancer cells from large volumes of liquid samples that can be translated to on-site clinic diagnosis.

Design and optimization of a novel bio-loom to weave melt-spun absorbable polymers for bone tissue engineering.

Bone graft procedures are currently among the most common surgical procedures performed worldwide, but due to high risk of complication and lack of viable donor tissue, there exists a need to develop alternatives for bone defect healing. Tissue engineering, for example, combining biocompatible scaffolds with mesenchymal stem cells to achieve new bone growth, is a possible solution. Recent work has highlighted the potential for woven polymer meshes to serve as bone tissue engineering scaffolds; since, scaffolds can be iteratively designed by adjusting weave settings, material types, and mesh parameters. However, there are a number of material and system challenges preventing the implementation of such a tissue engineering strategy. Fiber compliance, tensile strength, brittleness, cross-sectional geometry, and size present specific challenges for using traditional textile weaving methods. In the current work, two potential scaffold materials, melt-spun poly-l-lactide, and poly-l-lactide-co-ε-caprolactone, were investigated. An automated bio-loom was engineered and built to weave these materials. The bio-loom was used to successfully demonstrate the weaving of these difficult-to-handle fiber types into various mesh configurations and material combinations. The dobby-loom design, adapted with an air jet weft placement system, warp tension control system, and automated collection spool, provides minimal damage to the polymer fibers while overcoming the physical constraints presented by the inherent material structure. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 1342-1351, 2017.

Increased extracellular matrix density decreases MCF10A breast cell acinus formation in 3D culture conditions.

The extracellular matrix (ECM) contributes to the generation and dynamic of normal breast tissue, in particular to the generation of polarized acinar and ductal structures. In vitro 3D culture conditions, including variations in the composition of the ECM, have been shown to directly influence the formation and organization of acinus-like and duct-like structures. Furthermore, the density of the ECM appears to also play a role in the normal mammary tissue and tumour formation. Here we show that the density of the ECM directly influences the number, organization and function of breast acini. Briefly, non-malignant human breast MCF10A cells were incubated in increasing densities of a Matrigel®-collagen I matrix. Elastic moduli near and distant to the acinus structures were measured by atomic force microscopy, and the number of acinus structures was determined. Immunochemistry was used to investigate the expression levels of E-cadherin, laminin, matrix metalloproteinase-14 and ß-casein in MCF10A cells. The modulus of the ECM was significantly increased near the acinus structures and the number of acinus structures decreased with the increase in Matrigel-collagen I density. As evaluated by the expression of laminin, the organization of the acinus structures present was altered as the density of the ECM increased. Increases in both E-cadherin and MMP14 expression by MCF10A cells as ECM density increased were also observed. In contrast, MCF10A cells expressed lower ß-casein levels as the ECM density increased. Taken together, these observations highlight the key role of ECM density in modulating the number, organization and function of breast acini.

Remodeling of tannic acid crosslinked collagen type I induces apoptosis in ER+ breast cancer cells.

BACKGROUND: The naturally-occurring phytochemical tannic acid (TA) has anticancer properties. We have demonstrated that estrogen receptor-positive (ER+) breast cancer cells are more sensitive to effects of TA than triple-negative breast cancer cells and normal breast epithelial cells. In the present study, cells were grown on TA-crosslinked collagen beads. Growing cells remodel collagen and release TA, which affects attached cells. MATERIALS AND METHODS: The ER+ breast cancer cell line MCF7 and the normal breast epithelial cell line MCF10A were grown on TA-crosslinked collagen beads in roller bottles. Concentrations of TA in conditioned media were determined. Induced apoptosis was imaged and quantified. Caspase gene expression was calculated by real-time polymerase chain reaction (PCR). RESULTS: Both cell lines attached and grew on TA-crosslinked collagen beads where they remodeled collagen and released TA into surrounding medium. Released TA induced caspase-mediated apoptosis. CONCLUSION: TA induced apoptosis in a concentration-dependent manner, with ER+ MCF7 cells displaying more sensitivity to effects of TA.

The effect of wicking fibres in tissue-engineered bone scaffolds.

The major limitation of large tissue-engineered constructs used for bone regeneration is the lack of vasculature and, therefore, lack of transport of essential nutrients, chemical factors and progenitor cells. Research approaches to improve the transport properties of large scaffolds focus on using angiogenic factors and vasculogenic cells to create new vasculature; however, the slow rate of vessel formation and reliance on vessel self-assembly in these approaches is problematic. In this study, a novel approach has been proposed, using proprietary engineered 'wicking' fibres of non-circular cross-section that provide highly efficient transport for fluid and cells. The effect of wicking fibres on the movement of fluorescein isothiocyanate (FITC)-conjugated protein in a three-dimensional (3D) hydrogel system was analysed. The results indicated that the rate of diffusion of the fluorescent protein was greatly enhanced in hydrogels that contained wicking fibres in comparison to those that did not. The movement of progenitor cells along wicking fibres and round fibres was assessed. This study demonstrated that wicking fibres enhance the movement of critical growth factors and progenitor cells central for bone regeneration. The results suggested that the incorporation of wicking fibres into large tissue-engineered constructs may improve the transport of growth factors and progenitor cells essential for bone formation.

Simulator-based assessment of haptic surgical skill: a comparative study.

The aim of this study was to examine if the forces applied by users of a haptic simulator could be used to distinguish expert surgeons from novices. Seven surgeons with significant operating room expertise and 9 novices with no surgical experience participated in this study. The experimental task comprised exploring 4 virtual materials with the haptic device and learning the precise forces required to compress the materials to various depths. The virtual materials differed in their stiffness and force-displacement profiles. The results revealed that for nonlinear virtual materials, surgeons applied significantly greater magnitudes of force than novices. Furthermore, for the softer nonlinear and linear materials, surgeons were significantly more accurate in reproducing forces than novices. The results of this study suggest that the magnitudes of force measured using haptic simulators may be used to objectively differentiate experts' haptic skill from that of novices. This knowledge can inform the design of virtual reality surgical simulators and lead to the future incorporation of haptic skills training in medical school curricula.

Bone tissue engineering and regenerative medicine: targeting pathological fractures.

Patients with bone diseases have the highest risk of sustaining fractures and of suffering from nonunion bone healing due to tissue degeneration. Current fracture management strategies are limited in design and functionality and do not effectively promote bone healing within a diseased bone environment. Fracture management approaches include pharmaceutical therapy, surgical intervention, and tissue regeneration for fracture prevention, fracture stabilization, and fracture site regeneration, respectively. However, these strategies fail to accommodate the pathological nature of fragility fractures, leading to unwanted side effects, implant failures, and nonunions. To target fragility fractures, fracture management strategies should include bioactive bone substitutes designed for the pathological environment. However, the clinical outcome of these materials must be predictable within various disease environments. Initial development of a targeted treatment strategy should focus on simulating the physiological in vitro bone environment to predict clinical effectiveness of the engineered bone. An in vitro test system can facilitate reduction of implant failures and non-unions in fragility fractures.

Endovascular seldinger needle placement: a simulator for examining haptic skills.

In this work, we develop an affordable haptic simulator for examining haptic skills required for endovascular Seldinger needle placement.

A perspective on the role and utility of haptic feedback in laparoscopic skills training.

Laparoscopic surgery is a minimally invasive surgical technique with significant potential benefits to the patient, including shorter recovery time, less scarring, and decreased costs. There is a growing need to teach surgical trainees this emerging surgical technique. Simulators, ranging from simple "box" trainers to complex virtual reality (VR) trainers, have emerged as the most promising method for teaching basic laparoscopic surgical skills. Current box trainers require oversight from an expert surgeon for both training and assessing skills. VR trainers decrease the dependence on expert teachers during training by providing objective, real-time feedback and automatic skills evaluation. However, current VR trainers generally have limited credibility as a means to prepare new surgeons and have often fallen short of educators' expectations. Several researchers have speculated that the missing component in modern VR trainers is haptic feedback, which refers to the range of touch sensations encountered during surgery. These force types and ranges need to be adequately rendered by simulators for a more complete training experience. This article presents a perspective of the role and utility of haptic feedback during laparoscopic surgery and laparoscopic skills training by detailing the ranges and types of haptic sensations felt by the operating surgeon, along with quantitative studies of how this feedback is used. Further, a number of research studies that have documented human performance effects as a result of the presence of haptic feedback are critically reviewed. Finally, key research directions in using haptic feedback for laparoscopy training simulators are identified.

Role of vascularity for successful bone formation and repair.

Tissue engineering has been touted as the solution to regenerate tissue in patients. Yet current strategies for orthopedic application are limited because of the inability to successfully manage critical sized defects without a working vascular system. Bone grafts are commonly used in critical sized defects to fill the gap in missing bone tissue. Proper vasculature is vital to the success of these grafts to promote bone growth. The aim of this review is to describe the contribution of tissues surrounding critical sized defects, focusing in particular on the progenitor cell influx and factors contributing to neovascularization. An overview of clinical techniques to visualize patient vascular supply and evaluation of clinical techniques to increase blood flow to the critical defect site illustrates the current efforts of surgical intervention to promote proper bone formation. The opportunity and need lies in the development of tissue engineered bone grafts that can use and enhance available vascular supplies.

Tannic Acid preferentially targets estrogen receptor-positive breast cancer.

Research efforts investigating the potential of natural compounds in the fight against cancer are growing. Tannic acid (TA) belongs to the class of hydrolysable tannins and is found in numerous plants and foods. TA is a potent collagen cross-linking agent; the purpose of this study was to generate TA-cross-linked beads and assess the effects on breast cancer cell growth. Collagen beads were stable at body temperature following crosslinking. Exposure to collagen beads with higher levels of TA inhibited proliferation and induced apoptosis in normal and cancer cells. TA-induced apoptosis involved activation of caspase 3/7 and caspase 9 but not caspase 8. Breast cancer cells expressing the estrogen receptor were more susceptible to the effects of TA. Taken together the results suggest that TA has the potential to become an anti-ER(+) breast cancer treatment or preventative agent.

Evaluation of normal and metastatic mammary cells grown in different biomaterial matrices: establishing potential tissue test systems.

The in vitro growth and differentiation of normal mammalian cells is quite different than the growth of cells derived from tumors. Additionally, cells of the same origin (tissue) behave differently depending on the biomaterial matrix in or on which they are grown in vitro. We examined both Matrigel(TM) and a collagen/agarose blend and demonstrated that two murine mammary derived cells lines, 4T1 and NMuMG, derived from a metastatic mammary tumor or a normal mammary gland, respectively, exhibit different growth and differentiation patterns depending on the three-dimensional matrix in which they are grown. The shape and size of the colonies that formed were matrix dependent. The two cell lines produced different levels of growth factors and metalloproteinases, and expressed differentiation markers specific to a matrix. Through the classification of different cell behaviors in different growth matrices, we will be able to intelligently design and tune tissue test systems to ask and answer specific challenging scientific questions.

A haptic simulator to increase laparoscopic force application sensitivity.

Laparoscopic surgery demands perceptual-motor skills that are fundamentally different from open surgery, and laparoscopists must be adept at perceiving tissue interaction at the surgical site and then applying precise amounts of forces through instruments without damaging tissues. A haptic simulator that emulates multiple salient laparoscopic tasks and renders differing degrees of forces was created. Two of the haptic skills tasks were evaluated in two studies to determine their ability to distinguish and then train laparoscopic force application sensitivity. Results suggested that the simulator has the capability of rendering salient force feedback information to which novices become increasingly more perceptually sensitive.

Salient haptic skills trainer: initial validation of a novel simulator for training force-based laparoscopic surgical skills.

BACKGROUND: There is an increasing need for efficient training simulators to teach advanced laparoscopic skills beyond those imparted by a box trainer. In particular, force-based or haptic skills must be addressed in simulators, especially because a large percentage of surgical errors are caused by the over-application of force. In this work, the efficacy of a novel, salient haptic skills simulator is tested as a training tool for force-based laparoscopic skills. METHODS: Thirty novices with no previous laparoscopic experience trained on the simulator using a pre-test-feedback-post-test experiment model. Ten participants were randomly assigned to each of the three salient haptic skills-grasping, probing, and sweeping-on the simulator. Performance was assessed by comparing force performance metrics before and after training on the simulator. RESULTS: Data analysis indicated that absolute error decreased significantly for all three salient skills after training. Participants also generally decreased applied forces after training, especially at lower force levels. Overall, standard deviations also decreased after training, suggesting that participants improved their variability of applied forces. CONCLUSIONS: The novel, salient haptic skills simulator improved the precision and accuracy of participants when applying forces with the simulator. These results suggest that the simulator may be a viable tool for laparoscopic force skill training. However, further work must be undertaken to establish full validity. Nevertheless, this work presents important results toward addressing simulator-based force-skills training specifically and surgical skills training in general.

Overcoming hypoxia to improve tissue-engineering approaches to regenerative medicine.

The current clinical successes of tissue engineering are limited primarily to low-metabolism, acellular, pre-vascularized or thin tissues. Mass transport has been identified as the primary culprit, limiting the delivery of nutrients (such as oxygen and glucose) and removal of wastes, from tissues deep within a cellular scaffold. While strategies to develop sufficient vasculature to overcome hypoxia in vitro are promising, inconsistencies between the in vitro and the in vivo environments may still negate the effectiveness of large-volume tissue-engineered scaffolds. While a common theme in tissue engineering is to maximize oxygen supply, studies suggest that moderate oxygenation of cellular scaffolds during in vitro conditioning is preferable to high oxygen levels. Aiming for moderate oxygen values to prevent hypoxia while still promoting angiogenesis may be obtained by tailoring in vitro culture conditions to the oxygen environment the scaffold will experience upon implantation. This review discusses the causes and effects of tissue-engineering hypoxia and the optimization of oxygenation for the minimization of in vivo hypoxia.

Objective differentiation of force-based laparoscopic skills using a novel haptic simulator.

BACKGROUND: There is a growing need for effective surgical simulators to train the novice resident with a core skill set that can be later used in advanced operating room training. The most common simulator-based laparoscopic skills curriculum, the Fundamentals of Laparoscopic Skills (FLS), has been demonstrated to effectively teach basic surgical skills; however, a key deficiency in current surgical simulators is lack of validated training for force-based or haptic skills. In this study, a novel haptic simulator was examined for construct validity by determining its ability to differentiate between the force skills of surgeons and novices. METHODS: A total of 34 participants enrolled in the study and were divided into two groups: novices, with no previous surgical experience and surgeons, with some level of surgical experience (including upper level residents and attendings). All participants performed a force-based task using grasping, probing, or sweeping motions with laparoscopic tools on the simulator. In the first session, participants were given 3 trials to learn specific forces associated with locations on a graphic; after this, they were asked to reproduce forces at each of the locations in random order. A force-based metric (score) was used to record performance. RESULTS: On probing and grasping tasks, novices applied significantly greater overall forces than surgeons. When analyzed by force levels, novices applied greater forces on the probing task at lower and mid-range forces, for grasping at low-range forces ranges and, for sweeping at high-range forces. CONCLUSIONS: The haptic simulator successfully differentiated between novice and surgeon force skill level at specific ranges for all 3 salient haptic tasks, establishing initial construct validity of the haptic simulator. Based on these results, force-based simulator metrics may be used to objectively measure haptic skill level and potentially train residents. Haptic simulator development should focus on the 3 salient haptic skills (grasping, probing, and sweeping) where precise force application is necessary for successful task outcomes.

A quantitative metric for pattern fidelity of bioprinted cocultures.

This article describes a quantitative metric for coculture pattern fidelity and its use in the assessment of bioprinting systems. Increasingly, bioprinting is used to create in vitro cell and tissue models for the purpose of studying cell behavior and cell-cell interaction. To create meaningful models, a bioprinting system must be able to place cells in biologically relevant patterns with sufficient fidelity. A metric for assessing fidelity would be valuable for tuning experimental processes and parameters within a bioprinting system and for comparing performance between different systems. Toward this end, the "bioprinting fidelity index" (BFI), a metric which rates a bioprinted patterned coculture with a single number based on the proportions of correctly placed cells, is proposed. Additionally, a mathematical model of drop-on-demand printing is introduced, which predicts an upper bound on the BFI based on drop placement statistics. A proof-of-concept study was conducted in which patterned cocultures of D1 and 4T07 cells were produced in two different demonstration patterns. The BFI for the patterned cocultures was calculated and compared to the printing model fidelity prediction. The printing model successfully predicted the best BFI observed in the samples, and the BFI showed quantitatively that post-processing techniques negatively impacted the final fidelity of the samples. The BFI provides a principled method for comparing printing and post-processing methods.

Assessment of a Chitosan/Hyaluronan Injectable Composite for Fat Reconstruction.

The long-term success of autologous fat transplants is dependent on numerous factors including tissue quality, tissue survivability and the expansion of the implanted cells. The addition of a biomaterial filler to an injectable gel implant matrix provides an anchor and scaffold for proliferating cells as well as a carrier for syringe delivery. Building on the tissue reconstruction concept which we first disclosed, i.e., an injectable composite comprised of beads in a gel, the present study uses an injectable composite comprised of fatty-acid-loaded chitosan/gelatin (FA-CG) beads (i.e., a degradable filler material with tissue bulking potential) and hyaluronic acid (HA, a matrix). Human and bovine preadipocytes were considered independently; however, in both cases the cells proliferated and differentiated equally when grown with the FA-CG/HA composite, as demonstrated by the production of lipids and triglycerides, as well as expression of the human adipocyte marker aP2. The preadipocyte/FA-CG/HA injectables formed stable complexes that remained intact, with little degradation and no measurable immune response, for 4 months after implantation into mice. These results suggest that FA-CG/HA composite is a suitable injectable matrix for preadipocyte transplantation, providing the basis for further studies investigating the suitability of this technique for larger-scale tissue implantation and regeneration.

Characterizing the effects of cell settling on bioprinter output.

The time variation in bioprinter output, i.e. the number of cells per printed drop, was studied over the length of a typical printing experiment. This variation impacts the cell population size of bioprinted samples, which should ideally be consistent. The variation in output was specifically studied in the context of cell settling. The bioprinter studied is based on the thermal inkjet HP26A cartridge; however, the results are relevant to other cell delivery systems that draw fluid from a reservoir. A simple mathematical model suggests that the cell concentration in the bottom of the reservoir should increase linearly over time, up to some maximum, and that the cell output should be proportional to this concentration. Two studies were performed in which D1 murine stem cells and similarly sized polystyrene latex beads were printed. The bead output profiles were consistent with the model. The cell output profiles initially followed the increasing trend predicted by the settling model, but after several minutes the cell output peaked and then decreased. The decrease in cell output was found to be associated with the number of use cycles the cartridge had experienced. The differing results for beads and cells suggest that a biological process, such as adhesion, causes the decrease in cell output. Further work will be required to identify the exact process.