Novel Double Lumen Catheter for Drug Delivery at the Skin- Catheter Interface

This paper shows the technical feasibility of using a modified double lumen catheter for the delivery of fluids at the skin-catheter interface in an in vivo rabbit model. The modified catheter permits the extrusion of suspensions at the site of contact between the skin and the modified catheter (exit site). The modification consisted in the addition of a second lumen, exterior to the original one of the catheter and of shorter length, ending in an extrusion port placed sub dermally. An external fluid pump was attached to the catheter and the whole system was tested in vitro in human skin, in a cadaver model and in an in vivo rabbit model. Experiments were performed by delivering solutions of methylene blue in a controlled manner to the subcutaneous area to analyze the skin response to the process. Results indicate that this technique of pumping the material is effective in producing observable epithelial changes in the short term. The modification of the catheter addresses catheter exit site lack of integration of skin and works as a model that can be implemented in a number of percutaneous devices for long-term implantation.Medical Innovation Center of the University of MichiganPeer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/79405/1/peramo-catheter.pd

Characterization of a unique technique for culturing primary adult human epithelial progenitor/“stem cells”

Abstract Background Primary keratinocytes derived from epidermis, oral mucosa, and urothelium are used in construction of cell based wound healing devices and in regenerative medicine. This study presents in vitro technology that rapidly expands keratinocytes in culture by growing monolayers under large volumes of serum-free, essential fatty acid free, low calcium medium that is replaced every 24 hrs. Methods Primary cell cultures were produced from epidermal skin, oral mucosa and ureter by trypsinization of tissue. Cells were grown using Epilife medium with growth factors under high medium volumes. Once densely confluent, the keratinocyte monolayer produced cells in suspension in the overlying medium that can be harvested every 24 hrs. over a 7–10 day period. The cell suspension (approximately 8 X 105 cells/ml) is poured into a new flask to form another confluent monolayer over 2–4 days. This new culture, in turn produced additional cell suspensions that when serially passed expand the cell strain over 2–3 months, without the use of enzymes to split the cultures. The cell suspension, called epithelial Pop Up Keratinocytes (ePUKs) were analyzed for culture expansion, cell size and glucose utilization, attachment to carrier beads, micro-spheroid formation, induction of keratinocyte differentiation, and characterized by immunohistochemistry. Results The ePUKs expanded greatly in culture, attached to carrier beads, did not form micro-spheroids, used approximately 50% of medium glucose over 24 hrs., contained a greater portion of smaller diameter cells (8–10 microns), reverted to classical appearing cultures when returned to routine feeding schedules (48 hrs. and 15 ml/T-75 flask) and can be differentiated by either adding 1.2 mM medium calcium, or essential fatty acids. The ePUK cells are identified as cycling (Ki67 expressing) basal cells (p63, K14 expressing). Conclusions Using this primary culture technique, large quantities of epithelial cells can be generated without the use of the enzyme trypsin to split the cultures. The cells are small in diameter and have basal cell progenitor/”stem” (P/SC) cell characteristics induced by daily feeding with larger than normal medium volumes. The ePUK epithelial cells have the potential to be used in regenerative medicine and for basic studies of epithelia P/SC phenotype.http://deepblue.lib.umich.edu/bitstream/2027.42/112344/1/12895_2011_Article_127.pd

In Situ Polymerization of a Conductive Polymer in Acellular Muscle Tissue Constructs

We present a method to chemically deposit a conductive polymer, poly(3,4-ethylenedioxythiophene) (PEDOT), on acellularized muscle tissue constructs. Morphology and structure of the deposition was characterized using optical and scanning electron microscopies (SEM). The micrographs showed elongated, smooth, tubular PEDOT structures completely penetrating and surrounding the tissue fibers. The chemical polymerization was performed using iron chloride, a mild oxidizer. Remaining iron and chlorine in the tissue constructs were reduced to acceptable metabolic levels, while preserving the structural integrity of the tissue. We expect that these acellular, polymerized tissue implants will remain essentially unmodified in cellular environments in vitro and in vivo because of the chemical and thermal stability of the PEDOT polymer depositions. Our results indicate that in situ polymerization occurs throughout the tissue, converting it into an extensive acellular, non-antigenic substrate of interest for in vivo experiments related to nerve repair and bioartificial prosthesis. We expect these conducting polymer scaffolds to be useful for direct integration with electronically and ionically active tissues.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/63276/1/tea.2007.0123.pd

Tissue Engineering of Lips and Muco-Cutaneous Junctions: In Vitro Development of Tissue Engineered Constructs of Oral Mucosa and Skin for Lip Reconstruction

We report for the first time the fabrication of a three-dimensional tissue structure containing, in a continuous layer, the morphological features of a lip: epidermal skin, vermillion, and oral mucosa. This tissue engineered muco-cutaneous (M/C) equivalent was manufactured using human oral and skin keratinocytes grown on an acellular, nonimmunogenic dermal equivalent (AlloDerm-) to produce a tissue equivalent with similar anatomic and handling properties as native human lips. Confirmation of the structural composition of the construct was performed using routine histology and immunohistochemistry by identification of epithelial markers that are differentially expressed in separate anatomic areas of the lips. These full-thickness human lip skin equivalents can be used in surgical lip reconstruction in individuals suffering from lip loss from cancer, congenital deformations, and injuries after accidents. We propose this technique can be used as a general basis for tissue engineering of M/C junctions in other parts of the body, such as anus and vagina.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/90465/1/ten-2Etec-2E2011-2E0406.pd

Physical Characterization of Mouse Deep Vein Thrombosis Derived Microparticles by Differential Filtration with Nanopore Filters

With the objective of making advancements in the area of pro-thrombotic microparticle characterization in cardiovascular biology, we present a novel method to separate blood circulating microparticles using a membrane-based, nanopore filtration system. In this qualitative study, electron microscopy observations of these pro-thrombotic mouse microparticles, as well as mouse platelets and leukocytes obtained using a mouse inferior vena cava ligation model of deep-vein thrombosis are presented. In particular, we present mouse microparticle morphology and microstructure using SEM and TEM indicating that they appear to be mostly spherical with diameters in the 100 to 350 nm range. The nanopore filtration technique presented is focused on the development of novel methodologies to isolate and characterize blood circulating microparticles that can be used in conjunction with other methodologies. We believe that determination of microparticle size and structure is a critical step for the development of reliable assays with clinical or research application in thrombosis and it will contribute to the field of nanomedicine in thrombosis

Physical Studies of Glycosaminoglycans in Relation to the Adhesion Properties of Human Cancer Cells

The study of the processes relating glycobiology and cancer will have increased interest in coming years. To contribute to this trend the outcome of this work will be useful for investigations in glycobiology, using experimental methods exhibiting controlled carbohydrate composition, organization, and orientation, drawn from materials science and physics and that can be used in bioengineering and other technical areas in biology. In this work, the focus has been on physical studies of some members of the family of glycosaminoglycans and their role in cancer metastasis. The project studies the static adhesion of cancer cells to substrates functionalized with cell surface glycocalyx molecules and, in particular, in the interaction of heparan sulfate, keratan sulfate and chondroitin sulfates with the cells. Surface characterization techniques are used to analyze the structure of the polymeric brushes deposited on the substrates. The hypothesis that the adhesion of whole cancer cells to glysocaminoglycan substrates is a function of polysaccharide charge per dimer and chain length was proposed and tested.Part of the work has been dedicated to study the changes in the adhesion of tumor cells inthe presence of heparanase, an enzyme expressed in the tumor cell surface.The essential achievements of the project have been: a) Design of a new a method for the deposition and patterning of glycans to glass or silicon surfaces functionalized with a silane agent, exposing an amino terminated monolayer as functional substrate. b) Development of a new method for the calculation of the density of the deposited molecules. c) Physical characterization of the surfaces using a combination of surface science techniques, including ellipsometry and atomic force microscopy. These surfaces should be useful for developing additional experiments that may be helpful in understanding the adhesive properties of the cells. d) Comparative analysis of the behavior of cancer cells to the functionalized surfaces, specifically the study of the static adhesion of the cells, in the presence or absence of the surface protein heparanase or its inhibitors. e) Confirmation of the hypothesis that attachment of whole cancer cells, in vitro, depends linearly on the charge per dimer of polysaccharide

Solvated and generalised Born calculations differences using GPU CUDA and multi-CPU simulations of an antifreeze protein with AMBER

<p>While there has been an increase in the number of biomolecular computational studies employing graphics processing units (GPU), results describing their use with the molecular dynamics package AMBER with the CUDA implementation are scarce. No information is available comparing MD methodologies <i>pmemd.cuda, pmemd.mpi</i> or <i>sander.mpi,</i> available in AMBER, for generalised Born (GB) simulations or with solvated systems. As part of our current studies with antifreeze proteins (AFP), and for the previous reasons, we present details of our experience comparing performance of MD simulations at varied temperatures between multi-CPU runs using <i>sander.mpi, pmemd.mpi</i> and <i>pmemd.cuda</i> with the AFP from the fish ocean pout (1KDF). We found extremely small differences in total energies between multi-CPU and GPU CUDA implementations of AMBER12 in 1ns production simulations of the solvated system using the TIP3P water model. Additionally, GPU computations achieved typical one order of magnitude speedups when using mixed precision but were similar to CPU speeds when computing with double precision. However, we found that GB calculations were highly sensitive to the choice of initial GB parametrisation regardless of the type of methodology, with substantial differences in total energies.</p

Obra historica y panegyrica, escrita en obsequio del mayor de los nacidos, y Dios por autoridad con el mismo Dios, el gloriosissimo patriarca San Joseph

Sign.: ¶\p4\s, B-F\p4\s, A-Z\p4\s, 2A-2C\p4\s, 2D\p2\sPortada con orla tipográficaApostillas marginalesFrisos y viñetas xilográficasPort. con orla tipLa hoja de grabado calcográfico: "Benitez f." escudo nobiliarioLa segunda hoja de grabado calcográfico es una imagen de San JoséLa h. de grab. calc.: "Benitez f." escudo nobiliari

miRNAs in Tuberculosis: New Avenues for Diagnosis and Host-Directed Therapy

Tuberculosis (TB) is one of the most fatal infectious diseases and a leading cause of mortality, with 95% of these deaths occurring in developing countries. The causative agent, Mycobacterium tuberculosis (Mtb), has a well-established ability to circumvent the host’s immune system for its intracellular survival. microRNAs (miRNAs) are small, non-coding RNAs having an important function at the post-transcriptional level and are involved in shaping immunity by regulating the repertoire of genes expressed in immune cells. It has been established in recent studies that the innate immune response against TB is significantly regulated by miRNAs. Moreover, differential expression of miRNA in Mtb infection can reflect the disease progression and may help distinguish between active and latent TB infection (LTBI). These findings encouraged the application of miRNAs as potential biomarkers. Similarly, active participation of miRNAs in modulation of autophagy and apoptosis responses against Mtb opens an exciting avenue for the exploitation of miRNAs as host directed therapy (HDT) against TB. Nanoparticles mediated delivery of miRNAs to treat various diseases has been reported and this technology has a great potential to be used in TB. In reality, this exploitation of miRNAs as biomarkers and in HDT is still in its infancy stage, and more studies using animal models mimicking human TB are advocated to assess the role of miRNAs as biomarkers and therapeutic targets. In this review, we attempt to summarize the recent advancements in the role of miRNAs in TB as immune modulator, miRNAs’ capability to distinguish between active and latent TB and, finally, usage of miRNAs as therapeutic targets against TB