Papers by Keyword: Hepatocyte

Abstract: Since the promising virus -based gene therapies are often limited by problems such as the immunity of virus itself, the development of an efficient non-viral vector is of prime importance. For this reason, several synthetic nonviral polymeric carriers including cationic sequences have been molecularly designed. It is well known that the polymeric carriers with some cationic groups buffer the endosomal pH resulting in the enhanced transfection efficiency, but also in a relatively high toxicity. In the last decades, the polymers bearing pendant carbohydrates (glycopolymers) was proved to have relatively less toxic. Since the glycopolymers may not only decrease the toxicity of the cationic chain but also serve as targeting agent, we have rationally designed new glycopolymer-based gene delivery carriers. The interaction of carrier/gene polyplexes with hepatocytes and their intracellular trafficking were investigated in vitro. Our results show the significant efficacy of the galactose moieties on the uptake by hepatocytes, in a ligand specific manner.

Abstract: Recent advances in the preparation of nanofibre layers, especially using the Nanospider™ technology, allow prepare a sufficiently large area of nanofibrous layer of reproducible thickness and structure. Subsequently, it is possible to employ these layers as cell carriers and evaluate their efficiency in laboratory bioreactors. The construction of the functional hepatal bioreactor is particularly given by the positive response of hepatocyte cells to the used carrier layer as well as by the cell morphology, their viability and biological activity in certain period of time. We compared cell growth on collagen with nanofibres electrospun from selected copolymers of methacrylic esters (HEMA/EOEMA) and from differently prepared polycaprolactone (PCL) layers. The morphology was evaluated using Phaloidin/DAPI staining. On the nanofibres based on methacrylates, the cells survived and showed a common morphology comparing with cells grown on collagen (controls). On the PCL nanofibres, the cells attached well and showed a better growth than cells grown on collagen (controls). The results obtained in laboratory bioreactor proved the biochemical functionality of the studied system.

Abstract: The chitosan scaffold we prepared have a high porosity of about 90% with pore sizes from 50 to 200m. Lactose was conjugated onto the inner surface of the highly porous chitosan scaffold. It was used as substrate for rat hepatocytes culture. The cell attachment ratio was much higher than on monolayer membrane and non-modified porous scaffold. Metabolic activities of the cells were evaluated in terms of albumin secretion and urea synthesis. It was found that hepatocytes cultured on the modified scaffolds showed an increase in albumin secretion during the first 4 days and were more stable than that on non-modified scaffold. The results showed that the microstructure of porous scaffolds provides large surface for cells to adhere and facilitates nutrient and oxygen transportation. Such lactose modified scaffold has a potential application in bioartificial liver support system.

Abstract: Microcapsules of alginate cross-linked with divalent cations are the most common system for cell immobilization. In this work, the polyion complex (PIC) microcapsules were made using sodium alginate/barium chloride as the wall materials and gelatin/poly (vinyl alcohol) (PVA) as the extracellular matrices. The result of the permeability experiment of microcapsules using proteins with different molecular weight showed that the capsule has a molecular weight cut-off (MWCO) of 150 kDa. The hepatocytes encapsulated in microcapsules with gelatin and PVA in the core rapidly aggregated as incubation time increased. The aggregated hepatocytes showed high ammonia removal and albumin synthesis, showing a high potential for use in a bio artificial liver system.

Abstract: In this study, we prepared polystyrene (PS) nanofibers as hepatocytes culture substrates by electrospinning method and subsequently coated with specific ligand (poly(N-
-vinylbenzyl-- β-D-galactopyranosyl-(14)-D-gluconamide)(PVLA) for hepatocytes attachment. Rat hepatocytes’ behavior on the PVLA-coated and non-coated PS nanofibrous matrices have been investigated. Electrospun PS fiber structures revealed randomly aligned fibers with average diameter of 500 nm. Fabricated PS nanofibers had no bonding points like cotton fibers. Analyses by ATR/FTIR and ESCA revealed that PVLA was successfully coated to the surfaces of PS nanofibers. More hepatocytes were attached on the surface of PS nanofibers coated with PVLA than that on noncoated PS nanofibers. PS nanofibrous matrix could incorporate many cells into the interior of the matrix probably due to the suitable pore size. Cell viabilities cultured on PVLA-coated PS nanofibrous mats were maintained for 2 weeks, while it was decreased rapidly on PVLA-coated PS dishes. High hepatic function especially albumin secretion was maintained for 2 weeks on nanofibrous mats but rapidly decreased on flat PS dishes. These results indicate that nanofibrous structure enabled spheroid-like culture results in providing cell-cell communication and subsequent long-term maintenance of specific cell function.

Abstract: Micropatterned PEGylated substrates with two-dimensional arrays of plasma-etched circular domains (diameter:100 micro-m) were prepared by coating of mercapto-functionalized poly(ethylene glycol) (PEG) on Au surface, followed by plasma-etching through a metal mask pattern with circular holes. The PEGylated region on the patterned substrate works to repel proteins, consequently, inhibits cell adhesion. Then the micro-patterning of bovine articular chondrocytes or rat primary hepatocytes hetero-spheroids underlaid with human umbilical endothelial cells (HUVEC) was achieved on the plasma-etched circular domains, exposing the base gold surface. Obtained results suggested that the efficiency of inhibiting non-specific protein adsorption significantly affects on construction of micro-patterned cell adhesion and hetero-spheroids. The formation of hetero-spheroid thus suggested is significantly modulated by suface properties, particularly non-fouling character of PEG region. These arrayed spheroids is promising materials for tissue and cell-based biosensors (TBB/CBB) as well as tissue engineering technologies.

Abstract: To evaluate the biocompatibility of biomedical materials for hepatic tissue engineering, another new method was introduced to observe hepatocytes functions. In this experiment, hepatocytes were seeded onto four kinds of membranes of PLLA, PLGA (90:10), PLGA (75:25), and Chitosan cross-linked with collagen. The culture mediums were collected at 21 day after seeding, and then albumin (Alb), Urea (UN), glucose (Glu), total protein (TP), and triglyceride (TG) in the supernatant were detected by automatic biochemical analyzer. Results showed that hepatocytes on film of Chitosan cross-linked with collagen exhibited the highest level of TP and TG. These results were highly corresponding with the results of morphological observation. This data indicated that analyzing TP and TG in culture medium by automatic biochemical analyzer might be applied to evaluate hepatocytes biocompatibility on materials as a convenient and feasible method.

Abstract: Here, a two-dimensional microarray of ten thousand (100x100) hepatocyte hetero-spheroids, underlaid with endothelial cells, was successfully constructed with a 100 µm spacing in an active area of 20x20 mm on micro-fabricated glass substrates coated with poly(ethylene glycol) (PEG) brushes (Fig. 1). Co-cultivation of hepatocytes with endothelial cells was essential to stabilize hepatocyte viability and liver-specific functions, allowing us to obtain hepatocyte spheroids with a diameter of 100 µm, functioning as a miniaturized liver to secret albumin for at least three weeks. The spheroid array constructed here is highly useful as a platform of TBB and CBB to detect a wide variety of clinically, pharmacologically, and toxicologically active compounds through a cellular physiological response.

Abstract: To design the scaffold with suitable properties for the development of tissue engineered livers, materials design, selection and scaffold construct are three dispensable steps to be followed consequently. Firstly, some natural materials such as collagen, chitosan and alignate as well as some prevailing aliphatic polyester such as poly(lactic acid) (PLA), polyglycolide (PGA) and their copolymers poly(lactide-co-glycolide) (PLGA) are selected and characterized by hepatocyte culture. The experimental results reveal that the natural materials with excellent biocompatibility are not suitable as the scaffold alone because of the poor mechanical properties. At same time, aliphatic polyesters with good mechanical properties and biodegrade abilities are also proved inapplicable for hepatocytes for the lack of right cell recognition sites. Among our study scope, the hybrid materials such as collagen/chitosan or collagen/chitosan/heparin are the very promising candidates for hepatic tissue engineering scaffold. Secondly, two novel designs composed of collagen and PLGA, namely, the spindle porous scaffold with separated channels for transportation of nutrient, plasma and the one made by dewaxen-casting method are initiated respectively.