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Background: Glomerular endothelial cell (GEnC) fenestrations are recognised as an essential component of the glomerular filtration barrier, yet little is known about how they are regulated and their role in disease.Methods: We comprehensively characterized GEnC fenestral and functional renal filtration changes including measurement of glomerular ultrafiltration coefficient and glomerular filtration rate in diabetic mice (BTBR ob-/ob-). We also examined and compared human samples. We evaluated Eps homology domain protein-3 (EHD3) and its association with GEnC fenestrations in diabetes in disease samples and further explore its role as a potential regulator of fenestrations in an in vitro model of fenestration formation using b.End5 cells.Results: Loss of GEnC fenestration density was associated with decreased filtration function in diabetic nephropathy. We identified increased diaphragmed fenestrations in diabetes, which are posited to increase resistance to filtration and further contribute to decreased GFR. We identified decreased glomerular EHD3 expression in diabetes, which was significantly correlated with decreased fenestration density. Reduced fenestrations in EHD3 knock-down b.End5 cells in vitro further suggested a mechanistic role for EHD3 in fenestration formation.Conclusions: This study demonstrates the critical role of GEnC fenestrations in renal filtration function and suggests EHD3 may be a key regulator, loss of which may contribute to declining glomerular filtration function through aberrant GEnC fenestration regulation. This points to EHD3 as a novel therapeutic target to restore filtration function in disease

Infiltrative growth is a major cause of high lethality of malignant brain tumors such as glioblastoma (GBM). We show here that GBM cells upregulate guidance receptor Plexin-B2 to gain invasiveness. Deletion of Plexin-B2 in GBM stem cells limited tumor spread and shifted invasion paths from axon fiber tracts to perivascular routes. On a cellular level, Plexin-B2 adjusts cell adhesiveness, migratory responses to different matrix stiffness, and actomyosin dynamics, thus empowering GBM cells to leave stiff tumor bulk and infiltrate softer brain parenchyma. Correspondingly, gene signatures affected by Plexin-B2 were associated with locomotor regulation, matrix interactions, and cellular biomechanics. On a molecular level, the intracellular Ras-GAP domain contributed to Plexin-B2 function, while the signaling relationship with downstream effectors Rap1/2 appeared variable between GBM stem cell lines, reflecting intertumoral heterogeneity. Our studies establish Plexin-B2 as a modulator of cell biomechanics that is usurped by GBM cells to gain invasiveness., Huang et al demonstrate that glioblastoma cells upregulate axon guidance molecule Plexin-B2 to gain invasiveness and that Plexin-B2 promotes glioblastoma cell infiltration along axon fiber tracts in intracranial transplant models by modulating cellular biomechanics.

Multilayered skin substitutes comprising allogeneic cells have been tested for the treatment of nonhealing cutaneous ulcers. However, such nonnative skin grafts fail to permanently engraft because they lack dermal vascular networks important for integration with the host tissue. In this study, we describe the fabrication of an implantable multilayered vascularized bioengineered skin graft using 3D bioprinting. The graft is formed using one bioink containing human foreskin dermal fibroblasts (FBs), human endothelial cells (ECs) derived from cord blood human endothelial colony-forming cells (HECFCs), and human placental pericytes (PCs) suspended in rat tail type I collagen to form a dermis followed by printing with a second bioink containing human foreskin keratinocytes (KCs) to form an epidermis. In vitro, KCs replicate and mature to form a multilayered barrier, while the ECs and PCs self-assemble into interconnected microvascular networks. The PCs in the dermal bioink associate with EC-lined vascular structures and appear to improve KC maturation. When these 3D printed grafts are implanted on the dorsum of immunodeficient mice, the human EC-lined structures inosculate with mouse microvessels arising from the wound bed and become perfused within 4 weeks after implantation. The presence of PCs in the printed dermis enhances the invasion of the graft by host microvessels and the formation of an epidermal rete. Impact Statement Three Dimensional printing can be used to generate multilayered vascularized human skin grafts that can potentially overcome the limitations of graft survival observed in current avascular skin substitutes. Inclusion of human pericytes in the dermal bioink appears to improve both dermal and epidermal maturation.

Infiltrative growth is a major cause of the high lethality of malignant brain tumors such as glioblastoma (GBM). The study of the contribution of biomechanical processes to GBM invasion is an emerging field. We show here that GBM cells upregulate the guidance receptor Plexin-B2 to gain invasiveness by modulating their biomechanical properties. Deletion of Plexin-B2 in GBM stem cells limited tumor spread and shifted invasion paths from axon fiber tracts to perivascular routes. On a cellular level, Plexin-B2 adjusts cell adhesiveness, migratory responses to different matrix stiffness, and actomyosin dynamics, thus empowering GBM cells to leave stiff tumor bulk and infiltrate softer brain parenchyma. Correspondingly, gene signatures affected by Plexin-B2 were associated with locomotor regulation, matrix interactions, and cellular biomechanics. On a molecular level, the intracellular Ras-GAP domain contributed to Plexin-B2 function, while the signaling relationship with downstream effectors Rap1/2 appeared variable between GBM stem cell lines, reflecting intertumoral heterogeneity. Our studies have established Plexin-B2 as a modulator of cell biomechanics that is usurped by GBM cells to gain invasiveness. Ongoing investigations focus on the regulation of the biomechanical properties of cell membrane and cell actomyosin cortex through plexins that provide GBM cells with the mechanical dynamics to penetrate to restricted space.

Diffuse invasion of glioblastoma (GBM) cells into brain tissue is a key factor for its high lethality. GBM cell migration is affected by functions of plexins, which are transmembrane receptors of semaphorins that regulate cell adhesion and cytoskeletal dynamics. Expression of Plexin-B2 is upregulated in GBM and correlates with malignancy. We show here that Plexin-B2 activity regulates biomechanical properties of GBM cells, promoting invasive growth. Plexin-B2 activity increased the capacity of GBM to invade as dispersed single cells by reducing the cell-cell adhesiveness between GBM cells, indicating that a major function of Plexin-B2 activity is to downregulate cell-cell adhesion systems. RNA-Seq analyses also revealed that GBM stem cells (GSC) with deletion of Plexin-B2 altered expression of genes related to cell adhesion and the matrisome, indicating compensatory mechanisms in cellular dynamics. Interestingly, in vivo intracranial transplant studies demonstrated that growth and invasion of Plexin-B2 mutant GSC was impaired, with mutant cells invading shorter distances and migrating mainly as groups of cells forming chains. Plexin-B2 mutant cells also were more likely to adhere to the vasculature, rather than to fiber tracts, suggesting altered biomechanical properties. This shift may be related to high stiffness of basal lamina of the vasculature, as Plexin-B2 KO cells have a preference for migration on stiff substrate in vitro. Intriguingly, the loss in Plexin-B2 expression also changed the distribution of the mechanosensor transction factor YAP, with high expression of Plexin-B2 correlating with increased nuclear YAP. Structure-function analyses revealed that the Ras-GAP domain as main signaling output of Plexin-B2. The Rap proteins are pleiotropic regulators of cell adhesion and actomysosin contractility. Our data also showed that overexpression of Plexin-B2 can lead to decreased levels of Rap1/Rap2. Thus, Plexin-B2 acts as a key regulator of the adhesion and contractility of GBM cells, thereby facilitating their diffuse invasion.

Visualization is a valuable tool in problem solving, especially for citizen science games. In this study, we analyze data from 36,351 unique players of the citizen science game Foldit over a period of 5 years to understand how their choice of visualization options are affected by expertise and problem type. We identified clusters of visualization options, and found differences in how experts and novices view puzzles and that experts differentially change their views based on puzzle type. These results can inform new design approaches to help both novice and expert players visualize novel problems, develop expertise, and problem solve.

Glioblastoma multiforme (GBM) is an extremely lethal type of brain tumor as it frequently develops therapeutic resistance over months of chemotherapy cycles. Hence, there is a critical need to provide relevant biological systems to guide the development of new potent personalized drugs but also efficient methodologies that enable personalized prediction of various therapeutic regimens for enhanced patient prognosis. Towards this goal, we report on the development of i) an appropriate in vitro model that mimics the 3D tumor microenvironment and ii) a companion imaging modality that enables to assess this in vitro model in its entirety. More precisely, we developed an integrated platform of bio-printing in vitro 3D GBM models and mesoscopic imaging to monitor tumor growth and invasion along with long-term drug treatment. The newly-developed in vitro 3D model contains tumor spheroids made of patient-derived glioma stem cells with a fluorescent reporter and vascular channels for drug perfusion. The imaging of these thick tissue constructs was performed using our second-Generation Mesoscopic Fluorescence Molecular Tomography (2GMFMT) imaging system which delivered 3D reconstruction of the fluidic channels and the GBM spheroids over the course of pre- and post-drug treatment (up to 70 days). The 2D measurements collected via 2GMFMT was comparable to existing imaging modalities, but 2GMFMT enabled non-sacrificial volumetric monitoring that provided a unique insgiht into the GBM spheroid growth and drug response. Overall, our integrated platform provides customizable in vitro model systems combined with an efficient long-term non-sacrificial imaging for the volumetric change of tumor mass, thus has a great potential in profoundly affecting the drug pipeline for a vast array of pathologies as well as for guiding personalized therapeutic regimen.

Three-dimensional (3-D) cell printing, which can accurately deposit cells, biomaterial scaffolds and growth factors in precisely defined spatial patterns to form biomimetic tissue structures, has emerged as a powerful enabling technology to create live tissue and organ structures for drug discovery and tissue engineering applications. Unlike traditional 3-D printing that uses metals, plastics and polymers as the printing materials, cell printing has to be compatible with living cells and biological matrix. It is also required that the printing process preserves the biological functions of the cells and extracellular matrix, and to mimic the cell-matrix architectures and mechanical properties of the native tissues. Therefore, there are significant challenges in order to translate the technologies of traditional 3-D printing to cell printing, and ultimately achieve functional outcomes in the printed tissues. So it is essential to develop new technologies specially designed for cell printing and in-depth basic research in the bioprinted tissues, such as developing novel biomaterials specifically for cell printing applications, understanding the complex cell-matrix remodeling for the desired mechanical properties and functional outcomes, establishing proper vascular perfusion in bioprinted tissues, etc. In recent years, many exciting research progresses have been made in the 3-D cell printing technology and its application in engineering live tissue constructs. This review paper summarized the current development in 3-D cell printing technologies; focus on the outcomes of the live printed tissues and their potential applications in drug discovery and regenerative medicine. Current challenges and limitations are highlighted, and future directions of 3-D cell printing technology are also discussed.

Recent developments in three-dimensional (3D) bioprinting technology enable the efficient fabrication of thick and complex tissues. Numerous 3D-bioprinting techniques have been developed to create sophisticated structures in a wide range of scales (from micrometer to centimeter) with various biomaterials (Datta et al. 2017; Ozbolat et al. 2017). Unlike traditional 3D printing that prints inorganic materials and needs minimal postprocessing after the completion of printing, bioprinting involves more complicated components such as suitable cells and printable biomaterials, and also requires substantial postprinting procedures. To develop tissue-specific functions, long-term culture with appropriate stimuli is necessary for most of the engineered tissues. Vascularization of engineered tissues plays an important role in maintaining the tissue viability during the long-term postprinting maturation (Khademhosseini and Langer 2016; Kaully et al. 2009; Lovett et al. 2009; Phelps and Garcia 2010). Vascular networks with perfusion provide an adequate supply of nutrient and oxygen, thus maintaining viability and preventing necrosis of thick tissues during the maturation period. Vasculatures embedded within engineered tissues also provide backbone structures to recapitulate architectural features of the counterpart native tissue, and enable easy integration into the host circulatory system after the transplantation.

The success of engineered cell or tissue implants is dependent on vascular regeneration to meet adequate metabolic requirements. However, development of a broadly applicable strategy for stable and functional vascularization has remained challenging. We report here highly organized and resilient microvascular meshes fabricated through a controllable anchored self-assembly method. The microvascular meshes are scalable to centimeters, almost free of defects and transferrable to diverse substrates, ready for transplantation. They promote formation of functional blood vessels, with a density as high as ~220 vessels mm-2, in the poorly vascularized subcutaneous space of SCID-Beige mice. We further demonstrate the feasibility of fabricating microvascular meshes from human induced pluripotent stem cell-derived endothelial cells, opening a way to engineer patient-specific microvasculature. As a proof-of-concept for type 1 diabetes treatment, we combine microvascular meshes and subcutaneously transplanted rat islets and achieve correction of chemically induced diabetes in SCID-Beige mice for 3 months., The success of engineered tissue depends on the integration of a dense vascular network to supply nutrients and remove waste products. Here the authors design high density microvascular meshes made through an anchored self-assembly mechanism, and use these meshes to support subcutaneous pancreatic islet survival in a mouse diabetes model.

We developed a methodology using 3D bio-printing technology to create a functional in vitro vascular channel with perfused open lumen using only cells and biological matrices. The fabricated vasculature has a tight, confluent endothelium lining, presenting barrier function for both plasma protein and high-molecular weight dextran molecule. The fluidic vascular channel is capable of supporting the viability of tissue up to 5 mm in distance at 5 million cells/mL density under the physiological flow condition. In static-cultured vascular channels, active angiogenic sprouting from the vessel surface was observed whereas physiological flow strongly suppressed this process. Gene expression analysis was reported in this study to show the potential of this vessel model in vascular biology research. The methods have great potential in vascularized tissue fabrication using 3D bio-printing technology as the vascular channel is simultaneously created while cells and matrix are printed around the channel in desired 3D patterns. It can also serve as a unique experimental tool for investigating fundamental mechanisms of vascular remodeling with extracellular matrix and maturation process under 3D flow condition.

Although 3D bio-printing technology has great potential in creating complex tissues with multiple cell types and matrices, maintaining the viability of thick tissue construct for tissue growth and maturation after the printing is challenging due to lack of vascular perfusion. Perfused capillary network can be a solution for this issue; however, construction of a complete capillary network at single cell level using the existing technology is nearly impossible due to limitations in time and spatial resolution of the dispensing technology. To address the vascularization issue, we developed a 3D printing method to construct larger (lumen size of ~1 mm) fluidic vascular channels and to create adjacent capillary network through a natural maturation process, thus providing a feasible solution to connect the capillary network to the large perfused vascular channels. In our model, microvascular bed was formed in between two large fluidic vessels, and then connected to the vessels by angiogenic sprouting from the large channel edge. Our bio-printing technology has a great potential in engineering vascularized thick tissues and vascular niches, as the vascular channels are simultaneously created while cells and matrices are printed around the channels in desired 3D patterns.

Three-dimensional (3D) bioprinting, a flexible automated on-demand platform for the free-form fabrication of complex living architectures, is a novel approach for the design and engineering of human organs and tissues. Here, we demonstrate the potential of 3D bioprinting for tissue engineering using human skin as a prototypical example. Keratinocytes and fibroblasts were used as constituent cells to represent the epidermis and dermis, and collagen was used to represent the dermal matrix of the skin. Preliminary studies were conducted to optimize printing parameters for maximum cell viability as well as for the optimization of cell densities in the epidermis and dermis to mimic physiologically relevant attributes of human skin. Printed 3D constructs were cultured in submerged media conditions followed by exposure of the epidermal layer to the air-liquid interface to promote maturation and stratification. Histology and immunofluorescence characterization demonstrated that 3D printed skin tissue was morphologically and biologically representative of in vivo human skin tissue. In comparison with traditional methods for skin engineering, 3D bioprinting offers several advantages in terms of shape- and form retention, flexibility, reproducibility, and high culture throughput. It has a broad range of applications in transdermal and topical formulation discovery, dermal toxicity studies, and in designing autologous grafts for wound healing. The proof-of-concept studies presented here can be further extended for enhancing the complexity of the skin model via the incorporation of secondary and adnexal structures or the inclusion of diseased cells to serve as a model for studying the pathophysiology of skin diseases.

3D optical reconstruction of in-vitro Glioblastoma brain tumors was evaluated longitudinally with Mesoscopic Fluorescence Molecular Tomography. Tumor response to the Temozolomide, a clinical drug, were evaluated through volumetric change.

We present a direct cell printing technique to pattern neural cells in a three-dimensional (3D) multilayered collagen gel. A layer of collagen precursor was printed to provide a scaffold for the cells, and the rat embryonic neurons and astrocytes were subsequently printed on the layer. A solution of sodium bicarbonate was applied to the cell containing collagen layer as nebulized aerosols, which allowed the gelation of the collagen. This process was repeated layer-by-layer to construct the 3D cell-hydrogel composites. Upon characterizing the relationship between printing resolutions and the growth of printed neural cells, single/multiple layers of neural cell-hydrogel composites were constructed and cultured. The on-demand capability to print neural cells in a multilayered hydrogel scaffold offers flexibility in generating artificial 3D neural tissue composites.

We present a method to create multi-layered engineered tissue composites consisting of human skin fibroblasts and keratinocytes which mimic skin layers. Three-dimensional (3D) freeform fabrication (FF) technique, based on direct cell dispensing, was implemented using a robotic platform that prints collagen hydrogel precursor, fibroblasts and keratinocytes. A printed layer of cell-containing collagen was crosslinked by coating the layer with nebulized aqueous sodium bicarbonate. The process was repeated in layer-by-layer fashion on a planar tissue culture dish, resulting in two distinct cell layers of inner fibroblasts and outer keratinocytes. In order to demonstrate the ability to print and culture multi-layered cell-hydrogel composites on a non-planar surface for potential applications including skin wound repair, the technique was tested on a poly(dimethylsiloxane) (PDMS) mold with 3D surface contours as a target substrate. Highly viable proliferation of each cell layer was observed on both planar and non-planar surfaces. Our results suggest that organotypic skin tissue culture is feasible using on-demand cell printing technique with future potential application in creating skin grafts tailored for wound shape or artificial tissue assay for disease modeling and drug testing.

Objective Hyperglycemia worsens clinical outcomes in critically ill patients. Precise glycemia control using intravenous insulin improves outcomes. To determine if we could improve glycemia control over a previous paper-based, manual protocol, authors implemented, in a surgical intensive care unit (SICU), an intravenous insulin protocol integrated into a care provider order entry (CPOE) system. Design Retrospective before-after study of consecutive adult patients admitted to a SICU during pre (manual protocol, 32 days) and post (computer-based protocol, 49 days) periods. Measurements Percentage of glucose readings in ideal range of 70–109 mg/dl, and minutes spent in ideal range of control during the first 5 days of SICU stay. Results The computer-based protocol reduced time from first glucose measurement to initiation of insulin protocol, improved the percentage of all SICU glucose readings in the ideal range, and improved control in patients on IV insulin for ≥24 hours. Hypoglycemia ( Conclusion The CPOE-based intravenous insulin protocol improved glycemia control in SICU patients compared to a previous manual protocol, and reduced time to insulin therapy initiation. Integrating a computer-based insulin protocol into a CPOE system achieved efficient, safe, and effective glycemia control in SICU patients.

Arteries and veins have very different susceptibility to certain vascular diseases such as atherosclerosis and vascular calcification. The molecular mechanisms of these differences are not fully understood. In this study, we discovered that COUP-TFII, a transcription factor critical for establishing the venous identity during embryonic vascular development, also regulates the pathophysiological functions of adult blood vessels, especially those directly related to vascular diseases. Specifically, we found that suppression of COUP-TFII in venous ECs switched its phenotype toward pro-atherogenic by up-regulating the expression of inflammatory genes and down-regulating anti-thrombotic genes. ECs with COUP-TFII knockdown also readily undergo endothelial-to-mesenchymal transition (EndoMT) and subsequent osteogenic differentiation with dramatically increased osteogenic transcriptional program and calcium deposition. Consistently, over-expression of COUP-TFII led to the completely opposite effects. In vivo validation of these pro-atherogenic and osteogenic genes also demonstrates a broad consistent differential expression pattern in mouse aorta vs. vena cava ECs, which cannot be explained by the difference in hemodynamic flow. These data reveal phenotypic modulation by different levels of COUP-TFII in arterial and venous ECs and suggest COUP-TFII may play an important role in the different susceptibilities of arteries and veins to vascular diseases such as atherosclerosis and vascular calcification.

Glioblastoma multiforme (GBM), a malignant brain tumor, frequently exploit microvessels as guides for migration. Understanding cell-cell interaction between vascular cells and GBM cells may suggest a new therapeutic direction. We developed physiological 3-D glioma-vascular niche model to investigate this cell-cell interaction using 3-D bioprinting technology. In the model, patient-derived GBM cell cluster was closely located to fluidic vessel. The influence of microenvironmental factors (matrix composition) has been tested in order to provide better control on 3-D cell behavior in future research. The 3D vascular niche platform can be adapted to other biological systems and will be used as a valuable tool to model cell-cell interactions and to control microenvironment in other systems.

Three-dimensional (3D) bio-printing, a technology to create 3D tissue through layer-by-layer approach, offers great capacity to engineer tissue with desired cells, growth factors and biomaterial scaffolds in spatial patterns to mimic the native tissue architecture. With its flexibility and power, the 3D bio-printing technology can also be used to control stem cell fate and creating 3D stem cell niches. Meanwhile, 3D bio-printed tissues often incorporate thick opaque scaffold, dense population of cells, and are often large in size (1–100 mm). Thus, there are significant difficulties in visualizing the biological events within thick tissue constructs using current microscopic techniques. To elucidate the interaction of stem cells with the microenvironment in tissue engineering applications, it is necessary to develop novel molecular imaging techniques to non-invasively observe stem cell fate, cell-cell interactions, and structural features of an engineered tissue in real time. In this chapter, we review the usage of bio-printing technologies in stem cell and tissue engineering application, and the most recent development in the optical molecular imaging techniques for thick tissue imaging.

Due to diffusion limitation, tissues thicker than several hundred micrometers have difficulties in survival and proliferation. 3D bio-printing technology enables to create fluidic channels within tissue structures for appropriate media perfusion. This solution supports viability for the simple tissue model with limited number of cells and scaffold types; however, an additional capillary network is essential for more complex and denser tissues. In this study, we developed a 3D printing method to construct larger fluidic vascular channels in mm-scale and to create adjacent capillary network through a natural maturation process, thus providing a method to connect the capillary network to the large perfused vascular channels. Our bio-printing technology has a great potential in engineering vascularized thick tissues and vascular niches, as the vascular channels are simultaneously created while cells and matrices are printed around the channels in desired 3D patterns.

Melnick-Needles syndrome is an X-linked dominant bone dysplasia, lethal in males, characterized by a typical facies and characteristic radiological findings: including sclerosis of skull base and mastoids, S-shaped appearance of tibia; cortical irregularities with a ribbon appearance of the ribs. About 48 well-documented cases have been reported, most of them were sporadic. Parental transmission has been published in only 11 kindreds. We are presenting the first Brazilian family with mother-daughter transmission.The proposita presented the typical clinical and radiological features with characteristic facies, severe thoracic cage restriction and pulmonary hypertension. Her mother was more mildly affected.A síndrome de Melnick-Needles é uma displasia esquelética ligada ao X e letal no sexo masculino. Caracteriza-se pela presença de um fácies típico e dos seguintes achados radiológicos: esclerose dos ossos da base do crânio e mastóide, tíbia em forma de "S"; irregularidades corticais e costelas com aspecto de fita ("ribbon-like"). A maioria dos 48 casos já relatados na literatura (bem documentados), eram esporádicos, observando-se transmissão parental em apenas 11 famílias. Estudamos a primeira família brasileira com transmissão mãe-filha, cuja propósita apresentava o fácies e as manifestações radiológicas características da síndrome, além de uma restrição pulmonar importante e hipertensão pulmonar. Sua mãe, também afetada, mostrava um quadro clínico e radiológico mais leves.

Three-dimensional imaging of thick tissue constructs is one of the main challenges in the field of tissue engineering and regenerative medicine. Optical methods are the most promising as they offer noninvasive, fast, and inexpensive solutions. Herein, we report the use of mesoscopic fluorescence molecular tomography (MFMT) to image function and structure of thick bioprinted tissue hosted in a 3-mm-thick bioreactor. Collagen-based tissue assembled in this study contains two vascular channels formed by green fluorescent protein- and mCherry-expressing cells. Transfected live cell imaging enables us to image function, whereas Flash Red fluorescent bead perfusion into the vascular channel allows us to image structure. The MFMT optical reconstructions are benchmarked with classical microscopy techniques. MFMT and wide-field fluorescence microscopy data match within 92% in area and 84% in location, validating the accuracy of MFMT reconstructions. Our results demonstrate that MFMT is a well-suited imaging modality for fast, longitudinal, functional imaging of thick, and turbid tissue engineering constructs.

We report an application of Mesoscopic Fluorescence Molecular Tomography to 3D tissue engineering construct. Engineered thick tissue was hosting two 3D printed vasculatures. The channels were formed by live cells, expressing GFP and mCherry reporter genes, embedded in 3mm turbid media. Tissue and cells kept in a 3mm thick perfusion chamber during the entire imaging process which took less than 5 minutes.

Developing methods that provide adequate vascular perfusion is an important step toward engineering large functional tissues. Meanwhile, an imaging modality to assess the three-dimensional (3-D) structures and functions of the vascular channels is lacking for thick matrices (>2 ≈ 3 mm). Herein, we report on an original approach to construct and image 3-D dynamically perfused vascular structures in thick hydrogel scaffolds. In this work, we integrated a robotic 3-D cell printing technology with a mesoscopic fluorescence molecular tomography imaging system, and demonstrated the capability of the platform to construct perfused collagen scaffolds with endothelial lining and to image both the fluid flow and fluorescent-labeled living endothelial cells at high-frame rates, with high sensitivity and accuracy. These results establish the potential of integrating both 3-D cell printing and fluorescence mesoscopic imaging for functional and molecular studies in complex tissue-engineered tissues.

We integrated a mesoscopic fluorescence molecular tomography system, and visualized both the fluid flow and fluorescent-labeled living cells of vascular construct within thick scaffold at high-frame rates, with high sensitivity and accuracy.

We report on the development of a novel in vitro, multi-compartmental, three-dimensional, Human Skin Tissue Experimental Model (HISTEM) using three-dimensional free form fabrication (3DFFF). The availability of this model is expected to positively impact the development of transdermal vaccines and immunomodulators for infectious diseases and autoimmune disorders. Our studies demonstrate the biological and structural equivalence of HISTEM with ex vivo human skin.

In this work, we will report on the design and validation of a Laminar Optical Tomography systems developed at RPI to image tissue-engineered constructs. The systems characteristics and image reconstruction methods will be presented and discussed. In silico and in vitro validation of the imaging platform based on phantom studies mimicking the optical properties of various bio-tissues will demonstrate the potential of the system for tissue engineering applications, especially for imaging thick vascular constructs.

Sufficient vascularization of tissue is critical in regenerative medicine and tissue engineering. Embryonic stem cells (ESCs) can act as an unlimited resource in providing vascular tissue given their ability to self-renew and differentiate into any cell type. However, controlled vascular differentiation remains a challenge due to an intricate, coordinated interaction of multiple factors. Soluble factors, extracellular matrices, cell-cell interactions, and hemodynamic forces all contribute to vascular development. We propose to investigate how mouse ESCs respond to the combined stimuli of fluidic shear stress and various extracellular matrices, especially in terms of arterial or venous specification.

Vascularization for sufficient supplies of oxygen/nutrients and removal of waste in thicker tissue is one of the major challenges in tissue engineering. Here, we used three-dimensional bio-printing technology to engineer a vascular structure within hydrogel scaffold. Through layer-by-layer approach, we seeded endothelial cells in tubular form, which is embedded within three-dimensional collagen scaffold. Collagen hydrogel precursor was printed and polymerized. Endothelial cells in heated gelatin hydrogel were printed within the collagen scaffold, subsequently gelatin was liquefied and washed out in order to fabricate a fluidic channel with endothelial cell lining. The channels were connected to perfusion system. Specially-designed flow chamber was used for stable perfusion. The endothelial cells survived under flow condition, showing high viability and integration along the channel wall. The channel had a width 400–700 um, and was able to resist 200mmHg of pressure. Functional assessments of the printed vascular channels are in progress. We have demonstrated the capability of using three-dimensional bio-printing technology in rapid, simple, and highthroughput vasculature formation. The process has a potential application in vascular tissue engineering and study in angiogenesis.

Acetaminophen has unique analgesic and antipyretic properties. It is globally recommended as a first-line agent for the treatment of fever and pain due to its few contraindications. Acetaminophen lacks the significant gastrointestinal and cardiovascular side effects associated with nonsteroidal anti-inflammatory drugs and narcotics. An intravenous formulation of acetaminophen is available in Europe and is currently undergoing extensive clinical development for use in the United States. This use may have important implications for management of postoperative pain and fever. This review summarizes recent clinical trial experiences with intravenous acetaminophen for the treatment of postoperative pain and fever in adult and pediatric subjects.

One of the major challenges in tissue engineering is vascularization which provides adequate supplies of oxygen and nutrients to cells within thick tissue-engineered constructs. Oxygen, nutrients and other molecules required for cell growth and survival can only diffuse to 150∼200 μm without proper vascular system. For this reason, thicker tissues have diffusion problems and cannot survive/proliferate well. To date, fabrications of relatively thin tissues such as skin and bladder, and cartilage tissues, which require low level of oxygen and nutrients, are reported. Obstacles in vascularization still exist for thick and complex tissue construct such as kidney, lung and heart (1). Overcoming this problem is a critical step to the clinical applications of tissue engineering (2).Copyright © 2010 by ASME

Vascularization for adequate supplies of oxygen and nutrients is one of the challenges in tissue engineering. We report the three-dimensional cell printing technology to construct vascular structure in hydrogel scaffold which consist of endothelial cells and collagen scaffold. We constructed multi-layered, inter-connected channels within collagen hydrogel through layer-by-layer approach. Collagen hydrogel precursor was printed and cross-linked with a nebulized NaHCO 3 . Heated gelatin hydrogel was printed within the collagen scaffold as a sacrificial material, liquefied and washed out to fabricate channels after complete gelation of collagen scaffold. The channels was perfused by syringe pump showing ability to resist 200mmHg of trans-luminal pressure. Endothelial cells which form inner lining of the vascular system was printed within collagen precursor in tubular pattern. The capability of three-dimensional cell printing technology in vasculature formation without multiple complicated processes has a potential in vascular tissue engineering. The simplicity of the technology allows it to readily incorporate other types of cells or scaffold materials to construct complex tissue structure in the future.

One of the challenges in tissue engineering is to provide adequate supplies of oxygen and nutrients to cells within the engineered tissue construct. Soft-lithographic techniques have allowed the generation of hydrogel scaffolds containing a network of fluidic channels, but at the cost of complicated and often time-consuming manufacturing steps. We report a three-dimensional (3D) direct printing technique to construct hydrogel scaffolds containing fluidic channels. Cells can also be printed on to and embedded in the scaffold with this technique. Collagen hydrogel precursor was printed and subsequently crosslinked via nebulized sodium bicarbonate solution. A heated gelatin solution, which served as a sacrificial element for the fluidic channels, was printed between the collagen layers. The process was repeated layer-by-layer to form a 3D hydrogel block. The printed hydrogel block was heated to 37 degrees C, which allowed the gelatin to be selectively liquefied and drained, generating a hollow channel within the collagen scaffold. The dermal fibroblasts grown in a scaffold containing fluidic channels showed significantly elevated cell viability compared to the ones without any channels. The on-demand capability to print fluidic channel structures and cells in a 3D hydrogel scaffold offers flexibility in generating perfusable 3D artificial tissue composites.

Our earlier study indicated that functional magnetic resonance imaging (fMRI)-based detection and feedback of regional cortical activity from the auditory area enabled a group of individuals to increase the level of activation mediated by auditory attention during sound stimulation. The long-term ability to maintain an increased level of cortical activation, extending to a time period of a few weeks, however, has not been investigated. We used real-time fMRI to confirm the utility of fMRI in forming a basis for the regulation of brain function to increase the activation in the auditory areas, and demonstrated that the learned ability could be retained after a 2-week period, with additional involvement of an attention-related neural network.

This paper describes the application of an ensemble of indexing and classification systems, which have been shown to be successful in information retrieval and classification of medical literature, to a new task of assigning ICD-9-CM codes to the clinical history and impression sections of radiology reports. The basic methods used are: a modification of the NLM Medical Text Indexer system, SVM, k-NN and a simple pattern-matching method. The basic methods are combined using a variant of stacking. Evaluated in the context of a Medical NLP Challenge, fusion produced an F-score of 0.85 on the Challenge test set, which is considerably above the mean Challenge F-score of 0.77 for 44 participating groups.

Bioprinting technology has emerged as a powerful tool for building tissue and organ structures for drug discovery and regenerative medicine applications. In general, bioprint- ing uses a computer-controlled three-dimensional (3D) printing device to accurately deposit cells and biomaterials into precise geometries with the goal of creating anatomically correct biological structures. While traditional 3D printing uses metals, plastics, and polymers as printing materials or "ink", bioprinting deals with living cells and biological matrix. Hence, there are significant challenges to make a transition from traditional 3D printing to bioprint - ing, and ultimately achieve functional outcomes in bioprinted tissues. Therefore, it is critical that there is new technology development and in-depth basic research in bioprinted tissues, such as developing novel biomaterials specifically for use in bioprinting and biofabrication techniques, understanding the cell-matrix remodeling for the desired mechanical properties, and functional outcomes, establishing proper vascular perfusion, etc. Currently, there is active research going on bioprinting technology and its potential as a future source for tissue implants. This review paper overviews the current state of the art in bioprinting technology and focuses on the outcomes of the bioprinted tissues and their potential applications in drug discovery and regenerative medicine. Current challenges and limitations are highlighted, and future directions for next-generation bioprinting technology are also presented.