Increased curvature of hollow fiber membranes could up‐regulate differential functions of renal tubular cell layers

Tissue engineering devices as in vitro cell culture systems in scaffolds has encountered the bottleneck due to their much lower cell functions than real tissues/organs in vivo. Such situation has been improved in some extent by mimicking the cell microenvironments in vivo from either chemical or physical ways. However, microenvironmental curvature, commonly seen in real tissues/organs, has never been manipulated to regulate the cell performance in vitro. In this regard, this paper fabricated polysulfone membranes with or without polyethylene glycol modification to investigate the impact of curvature on two renal tubular cells. Regardless the varying membrane curvatures among hollow fiber membranes of different diameters and flat membrane of zero curvature, both renal cells could well attach at 4 h of seeding and form similar confluent layers at 6 days on each membrane. Nevertheless, the renal cells on hollow fibers, though showing confluent morphology as those on flat membranes, expressed higher renal functions and, moreover, the renal functions significantly increased with the membrane curvature among hollow fibers. Such upregulation on functions was unassociated with mass transport barrier of hollow fibers, because the cultures on lengthwise cut hollow fibers without mass transfer barrier showed same curvature effect on renal functions as whole hollow fibers. It could be proposed that the curvature of hollow fiber membrane approaching to the large curvature in kidney tubules increased the mechanical stress in the renal cells and thus might up‐regulate the renal cell functions. In conclusion, the increase of substrate curvature could up‐regulate the cell functions without altering the confluent cell morphology and this finding will facilitate the design of functional tissue engineering devices. Biotechnol. Bioeng. 2013; 110: 2173–2183. © 2013 Wiley Periodicals, Inc.     

A radial flow hollow fiber bioreactor for the large-scale culture of mammalian cells

A radial flow hollow fiber bioreactor has been developed that maximizes the utilization of fiber surface for cell growth while eliminating nutrient and metabolic gradients inherent in conventional hollow fiber cartridges. The reactor consists of a central flow distributor tube surrounded by an annular bed of hollow fibers. The central flow distributor tube ensures an axially uniform radial convective flow of nutrients across the fiber bed. Cells attach and proliferate on the outer surface of the fibers. The fibers are pretreated with polylysine to facilitate cell attachment and long-term maintenance of tissuelike densities of cell mass. A mixture of air and CO(2) is fed through the tube side of the hollow fibers, ensuring direct oxygenation of the cells and maintenance of pH. Spent medium diffuses across the cell layer into the tube side of the fibers and is convected away along with the spent gas stream. The bioreactor was run as a recycle reactor to permit maximum utilization of nutrient medium. A bioreactor with a membrane surface area of 1150 cm(2) was developed and H1 cells were grown to a density of 7.3 x 10(6) cells/cm(2).     

Rapid deoxygenation of red cells and hemoglobin solution using hollow capillary fibers

A newly designed capillary deoxygenator has been constructed by using microporous polypropylene hollow fibers sealed into an airtight plexiglass housing. Oxygenated red cell suspensions and hemoglobin solutions flowing through the hollow fibers were subjected to deoxygenation with a gas mixture composed of 95 percent N2 and 5 percent CO2 passed through the housing. At a given flow rate of the oxygenated fluid, the outgoing fluid pO2 varied directly with hematocrit and inversely with the residence time. With a deoxygenator composed of 144 parallel 100-micrometers fibers with an active length of 10 cm, 2 ml of blood at 10 percent hematocrit can be converted from arterial to venous pO2 in approximately 1 min. The design of this deoxygenator provides a method for rapid deoxygenation of blood without red cell membrane damage or hemolysis.     

Arachidonic acid-rich oil production by Mortierella alpina with different gas distributors

Arachidonic acid (ARA)-rich oil production by Mortierella alpina is a high oxygen demand and shear-sensitive process. In the aerobic fermentation process, oxygen supply is usually a limiting factor owing to the low solubility of oxygen in the fermentation broth. Two kinds of perforated ring gas distributors and a novel microporous ceramic membrane gas distributor were designed and applied to improve oxygen supply. With the decrease of the orifice diameter of perforated ring gas distributors, dry cell weight (DCW), lipids concentration, and ARA content in total fatty acid increased from 17.86 g/L, 7.08 g/L, and 28.08 % to 25.67 g/L, 11.94 g/L, and 36.99 %, respectively. Furthermore, the effect of different dissolved oxygen (DO) on ARA-rich oil production with membrane gas distributor was also studied. The maximum DCW, lipid concentration, and ARA content using membrane gas distributor with DO controlled at 40 % reached 29.67 g/L, 16.74 g/L, and 49.53 %, respectively. The ARA titer increased from 1.99 to 8.29 g/L using the membrane gas distributor to substitute the perforated ring gas distributor. In the further experiment, a novel tubular titanium metal membrane gas distributor was successfully applied in a 7,000 L bioreactor and the results demonstrated that membrane gas distributor was industrially practical.     

Treatment of trichloroethylene (TCE) in a membrane biofilter

This article reports on the biodegradation of trichloroethylene (TCE) in a hollow-fiber membrane biofilter. Air contaminated with TCE was passed through microporous hollow fibers while an oxygen-free nutrient solution was recirculated through the shell side of the membrane module. The biomass was attached to the outside surface of the microporous hollow fibers by initially supplying toluene in the gas phase that flows through the fibers. While studies on TCE biodegradation were conducted, there was no toluene present in the gas phase. At 20-ppmv inlet concentration of TCE and 36-s gas-phase residence time, based on total internal volume of the hollow fibers, 30% removal efficiency of TCE was attained. At higher air flow rates or lower gas-phase residence times, lower removal efficiencies were observed. During TCE degradation, the pH of the liquid phase on the shell side of the membrane module decreased due to release of chloride ions. A mathematical model was developed to describe the synchronous aerobic/anaerobic biodegradation of TCE. (c) 1996 John Wiley & Sons, Inc.     

Mass transfer in blood oxygenators using blood analogue fluids

Mass transfer correlations for hollow fiber blood oxygenators have been determined experimentally using Newtonian and non-Newtonian blood analogue fluids. The Newtonian fluids consisted of deionized water and glycerol/water mixtures. The non-Newtonian fluids were prepared by adding small amounts of xanthan gum to the Newtonian blood analogue fluids. The rheological behavior of the non-Newtonian blood analogue fluids was modeled using the power law. The diffusion of oxygen into and out of the Newtonian and non-Newtonian blood analogue fluids has been studied. The liquid stream flowed outside and across bundles of woven hollow fibers, while the gas stream flowed inside the fibers.     

Membranes and bioreactors: a technical challenge in biotechnology

Integrating the properties of synthetic membranes with biological catalysts such as cells and enzymes forms the basis of an exciting new technology called membrane bioreactors. The impetus behind this marriage comes from the recent spectacular advances in recombinant DMA and cell fusion technologies and the need to develop competitive bioprocessing schemes to produce complex and active biological molecules. The advantages and limitations of using membrane bioreactors for entrapping whole cells and enzymes are reviewed. Various membrane configurations such as microcapsules, hollow fibers, and flat sheets are compared. Several different entrapped membrane bioreactors, including single, laminated and microporous, for the conversion of optically active enantiomers are described. As with new and exciting technologies, the future of membrane bioreactors in biotechnology will depend on their ability to produce desired molecules at competitive costs.     

CH4 production from H2 and CO2 byMethanobacterium thermoautotrophicum cells fixed on hollow fibers

Summary Methane was produced from H₂ and CO₂ byMethanobacterium thermoautotrophicum cells fixed on the surface of hollow fibers. The mineral solution permeated through the inside of fibers was consumed by the cells, while the gaseous substrate flowing outside the fibers was directly metabolized to methane. Methane production was proportional to hollow fiber length i.e., contact area between cell layer and gas phase. In repeated batch cultures, the production rates of methane and cell mass were 33.1 L/L reactor/day and 1.75 g cells/L reactor/day, respectively with 90% conversion rate.     

Culture of primary rat hepatocytes within a flat hollow fibre cassette for potential use as a component of a bioartificial liver support system

Primary rat hepatocytes were cultured in a flat, hollow-fibre cassette, `The Tecnomouse', which provided direct oxygenation and a homogeneous environment for cells within the cassette. Most hollow fibre systems utilise media oxygenators to provide O₂ to cells; in the Tecnomouse cassette, cells are provided with direct oxygenation via gas channels in the silicone membrane surrounding the hollow fibres. Hepatocyte functionality was monitored by following urea production, albumin production and cytochrome P-450 enzyme activities. The system could maintain cells in a viable state and the presence of specific hepatocyte functions including albumin production and cytochrome P-450 activity. Electron microscopy showed aggregated spherical hepatocytes and apparent high extent of necrosis.     

Kinetics studies in a continuous steady state hollow fiber membrane enzyme reactor

Porous hollow cellulose fibers have been used to separate a nonflowing enzyme solution of alkaline phosphatase from a continuous flow of substrate. The porosity of the hollow fiber membrane allows the substrate and product to diffuse freely through the membrane while restricting the permeation of the enzyme. The resulting “immobilized” enzyme system has been shown to behave as a continuous reactor—converting p-nitrophenylphosphate to p-nitrophenol. By varying the concentrations, flow rate, etc., either diffusion or enzyme kinetics can be studied. The continual influx of product and removal of substrate at steady state allows the study of kinetics of relatively short half-life enzymes and unstable systems.     

A numerical evaluation of a hollow fiber extractive fermentor process for the production of ethanol

The economics of a process for the production of ethanol employing a hollow fiber extractive fermentor have been investigated. A computer simulation of the process incorporating a mathematical model of the fermentor was used to calculate the mass and energy balances. The results of the process simulation were read into a computer spreadsheet programmed with the economic calculations from which a final ethanol product cost was obtained. The process was found to be as competitive as conventional fermentation processes even at the currently high cost--$4/sq ft--of hollow fibers. It was determined that the 1986 price of 46.2 cents/L of ethanol produced by the process would be reduced by 1.8 cents/L for every $1/sq foot drop in the price of hollow fibers. A comparison of this process with conventional fermentation processes indicates that its potential savings lie in its ability to use a concentrated sugar feed, and the fermentor's increased productivity and ability to produce a concentrated ethanol stream which is removed by the extracting solvent.     

Immobilized enzyme cellulose hollow fibers: III. Physical properties and in vitro biocompatibility

The initial testing of the safety of a cellulose-heparinase hollow fiber device was assessed with respect to physical properties and in vitro biocompatibility. The material cleared urea and creatinine without passing albumin, even at high flow rates. The clearance of urea and creatinine by cellulose-heparinase was equal or slightly reduced in comparision to the cellulose device. The cellulose-neparinase device tolerance to now rates was also unchanged. In addition, scanning electron microscopy of the lumen established the uniformity of the material. The analysis of clearance rates and the scanning electron micrographs show there to be no damage to the cellulose membrane after tresyl chloride activation and heparinase immobilization. The investigation of biocompatibility in an in vitro test system with whole human blood indicated that there were no significant changes in the biocompatibility of cellulose with bound heparinase. There was no change in the level of red blood cells, white blood cells, or platelets over the course of in vitro whole blood perfusion through cellulose or cellulose-heparinase hollow fiber devices. Low levels of plasma hemoglobin and complement activation were observed with cellulose and cellulose-heparinase devices. Thus, the cellulose hollow fibers can be functionalized without any changes in in vitro performance.     

Modulation of transmembrane pressure in manufacturing scale tangential flow filtration N-1 perfusion seed culture

Mammalian cells were grown to high density in a 3,000 L culture using perfusion with hollow fibers operated in a tangential flow filtration mode. The high-density culture was used to inoculate the production stage of a biomanufacturing process. At constant permeate flux operation, increased transmembrane pressures (TMPs) were observed on the final day of the manufacturing batches. Small scale studies suggested that the filters were not irreversibly fouled, but rather exposed to membrane concentration polarization that could be relieved by tangential sweeping of the hollow fibers. Studies were undertaken to analyze parameters that influence the hydrodynamic profile within hollow fibers; including filter area, cell density, recirculation flow rate, and permeate flow rate. Results indicated that permeate flow rate had the greatest influence on modulating TMP. Further evaluation showed a significant decrease in TMP when permeate flow was reduced, and this occurred without any negative effect on cell growth or viability. Hence, a 30% reduction of permeate flow rate was implemented at manufacturing scale. A stable operation was achieved as TMP was successfully reduced by 75% while preserving all critical factors for performance in the perfusion bioreactor.     

Hollow fiber culture accelerates differentiation of Caco-2 cells

Caco-2 cells usually require 21 days of culture for developing sufficient differentiation in traditional two-dimensional Transwell culture, deviating far away from the quick differentiation of enterocytes in vivo. The recently proposed three-dimensional cultures of Caco-2 cells, though imitating the villi/crypt-like microstructure of intestinal epithelium, showed no effect on accelerating the differentiation of Caco-2 cells. In this study, a novel culture of Caco-2 cells on hollow fiber bioreactor was applied to morphologically mimic the human small intestine lumen for accelerating the expression of intestine functions. The porous hollow fibers of polyethersulfone (PES), a suitable membrane material for Caco-2 cell culture, successfully promoted cells to form confluent monolayer on the inner surface. The differentiated functions of Caco-2 cells, represented by alkaline phosphatase, γ-glutamyltransferase, and P-glycoprotein activity, were greatly higher in a 10-day hollow fiber culture than in a 21-day Transwell culture. Moreover, the Caco-2 cells on PES hollow fibers expressed higher F-actin and zonula occludens-1 protein than those on Transwell culture, indicative of an increased mechanical stress in Caco-2 cells on PES hollow fibers. The accelerated differentiation of Caco-2 cells on PES hollow fibers was unassociated with membrane chemical composition and surface roughness, but could be stimulated by hollow fiber configuration, since PES flat membranes with either rough or smooth surface failed to enhance the differentiation of Caco-2. Therefore, the accelerated expression of Caco-2 cell function on hollow fiber culture might show great values in simulation of the tissue microenvironment in vivo and guide the construction of intestinal tissue engineering apparatus.     

Slit-Surface Electrospinning: A Novel Process Developed for High-Throughput Fabrication of Core-Sheath Fibers

In this work, we report on the development of slit-surface electrospinning – a process that co-localizes two solutions along a slit surface to spontaneously emit multiple core-sheath cone-jets at rates of up to 1 L/h. To the best of our knowledge, this is the first time that production of electrospun core-sheath fibers has been scaled to this magnitude. Fibers produced in this study were defect-free (i.e. non-beaded) and core-sheath geometry was visually confirmed under scanning electron microscopy. The versatility of our system was demonstrated by fabrication of (1) fibers encapsulating a drug, (2) bicomponent fibers, (3) hollow fibers, and (4) fibers from a polymer that is not normally electrospinnable. Additionally, we demonstrate control of the process by modulating parameters such as flow rate, solution viscosity, and fixture design. The technological achievements demonstrated in this work significantly advance core-sheath electrospinning towards commercial and manufacturing viability.     

A new coiled hollow-fiber module design for enhanced microfiltration performance in biotechnology.

The microfiltration performance of a novel membrane module design with helically wound hollow fibers is compared with that obtained with a standard commercial-type crossflow module containing linear hollow fibers. Cell suspensions (yeast, E. coli, and mammalian cell cultures) commonly clarified in the biotechnology industry are used for this comparison. The effect of variables such as transmembrane pressure, particle suspension concentration, and feed flow rate on membrane performance is evaluated. Normalized permeation fluxes versus flow rate or Dean number behave according to a heat transfer correlation obtained with centrifugal instabilities of the Taylor type. The microfiltration performance of this new module design, which uses secondary flows in helical tubes, is significantly better than an equivalent current commercial crossflow module when filtering suspensions relevant to the biotechnology industry. Flux and capacity improvements of up to 3.2-fold (constant transmembrane pressure operation) and 3.9-fold (constant flux operation), respectively, were obtained with the helical module over those for the linear module.     

Fine-structural and morphometrical studies on the segmental septa of the giant fibers in the earthworm,Eisenia foetida

Summary The fine structure of junctional specializations on the segmental septa in the median and lateral giant fibers of the earthworm (Eisenia foetida) was examined. Eight morphologically different septal domains were identified; a gap junction, a junction with hemispherical hollow structures, a chemical synapse-like junction, intermediate type and punctum adherens type junctions, a junction with adjoining vesicular layers, an area flanked by flattened membranous sacs, a non specialized area, and an area consisting of widely separated membranes with interposed glial processes. The area of each domain was measured by a cytometrical technique using quasi-serial sections. The gap junction occupied 3% and 0.2% of the septal area of the median and lateral giant fibers, respectively. Junctions with hemisperical hollow structures, characteristic of the earthworm giant fibers, occupied 2.5% and 13.9% of the median and lateral giant fibers, respectively. Various membrane domains except the gap junction, the junction with hemispherical hollow structures, and the chemical synapse-like junction accounted for similar proportional areas in both median and lateral giant fibers.The functional implications of these junctional specializations, especially the gap junction, are discussed.     

Theoretical analysis of the effect of convective flow on solute transport and insulin release in a hollow fiber bioartificial pancreas

The bioartificial pancreas, in which transplanted pancreatic tissue or isolated cells are cultured on a hollow fiber membrane, is an attractive approach to restore physiologic insulin delivery in the treatment of diabetes. Insulin response in prototype devices has been unacceptable due to the large mass transport limitations associated with the membrane and the surrounding shell region. Although available theoretical analyses provide some insight into the combined effects of transport and reaction in the bioartificial pancreas, they cannot quantitatively account for the effects of convective recirculation flow, complex intrinsic insulin secretory kinetics, and non-uniform distribution of pancreatic cells. We have developed a detailed model for glucose and insulin transport and insulin secretion in the hollow fiber bioartificial pancreas based on the solution of the mass and momentum conservation equations describing flow and transport in the lumen, matrix, and shell. Model predictions are in good agreement with literature data obtained in a hollow fiber device with minimal radial convective flow. Although no quantitative data are available for a device with significant radial convection, model simulations demonstrate that convective recirculation flow can dramatically improve insulin response, allowing the device to accurately capture the bi-phasic insulin secretion characteristic of the normal physiologic response. Results provide fundamental insights into the coupling between kinetics and transport in the hollow fiber system and a rational basis for the design of clinical devices.     

Study of banana and coconut fibers Botanical composition, thermal degradation and textural observations

Four fibers from banana-trees (leaf, trunk) and coconut-tree (husk, fabric) were examined before their incorporation in cementitious matrices, in order to prepare insulating material for construction. Their botanical compositions have been determined following a method described previously. Thermal degradation of these fibers were studied between 200 and 700 degrees C under nitrogen gas flow. Temperature of pyrolysis was the experimental parameter investigated. The solid residues obtained were analyzed by classical elemental analysis, Fourier Transform Infra Red (FTIR) spectroscopy and were observed by Scanning Electron Microscopy (SEM). This study has shown (1) the relation between botanical, chemical composition with both localization of fibers in the tree and type of tree; (2) the rapid and preferential decomposition of banana fibers with increasing temperature of pyrolysis and (3) the rough samples are made of hollow fibers.     

Mathematical model of microporous hollow-fiber membrane extractive fermentor

A mathematical model is presented for a microporous hollow-fiber membrane extractive fermentor (HFEF). The model is based on the continuous flow of the aqueous nutrient phase and cells through the shell space of the fermentor where the fermentation reaction occurs. The product diffuses from the shell space through the hollow-fiber membrane where it is continuously removed by solvent flowing concurrently through the fiber lumen. Results for ethanol production show that the HFEF has a volumetric productivity significantly higher than that possible using conventional methods. The model predicts the existence of an optimum volume fraction of hollow fibers in the fermentor that maximizes the total volumetric productivity. This optimum is the result of a classic trade-off between the volume fraction of the fermentor required for fermentation and that required for efficient removal of the ethanol product to minimize product inhibition.