To investigate differentiation and growth process of keratinocytes in organotypic cultured skin, we carried out immunohistochemical studies for cytokeratin (CK) 10, 14, 16, 17 and involucrin in the cultured skin. In normal skin CK14 and CK10 were detected in the basal and all suprabasal layer, respectively, whereas in artificial skin CK14 was detected up to the middle of spinous layer but CK10 expressed from the middle of spinous layer. The detection of involucrin in normal skin was from the upper spinous layer but found from lower spinous layer in the artificial skin. Both CK16 and CK17 did not expressed in in vivo skin but expressed weakly in the spinous layer of artificial skin. It is therefore concluded that the characteristics of basal cell were maintained in the several, lower layers of the sartificial skin. The growth and differentiation steps of the skin were similar to those of in vivo although differences were seen in the expression level.
Cells, Cultured* ; Epidermis* ; Immunohistochemistry ; Keratinocytes ; Keratins* ; Skin ; Skin, Artificial*

The temporary function replacement and intensive support play a pivotal role in the treatment of patients with acute or chronic end-stage organ failure. The hemodialysis and peritoneal dialysis have become routine techniques in the management of acute or chronic renal failure. But for the complexity of hepatic function, e.g. metabolism, biosynthesis and biotransformation for endogenous or exogenous substrates, the simulation or partial replacement of hepatic function is a great dream of bionic technologists. With the development of cell biology and biomedical material, the combination of biomaterial and the hepatocyte cultured ex vivo could provide a range of human liver-specific functions. The combination of biomaterial and viable cell was called hybrid or bioartificial liver support system (BALSS). It is a chimera of biomaterials and hepatocytes. The bionic technologists study the cell and its culture in vitro, which is the main component of BASLL. Many types of BALSS were translated into the early clinical stage. In this overview, we review the hepatocyte culture and the design of different bioreactors. It includes the immune obstacles in xeno-hemoperfusion and how to assess pre-clinical and clinical effectiveness.
Bioreactors ; Cells, Cultured ; Hepatocytes ; cytology ; Humans ; Liver, Artificial

It is clear that the construction of large insert DNA libraries is important for map-based gene cloning, the assembly of physical maps, and simple screening for specific genomic sequences. The bacterial artificial chromosome (BAC) system is likely to be an important tool for map-based cloning of genes since BAC libraries can be constructed simply and analyzed more efficiently than yeast artificial chromosome (YAC) libraries. BACs have significantly expanded the size of fragments from eukaryotic genomes that can be cloned in Escherichia coli as plasmid molecules. To facilitate the isolation of molecular-biologically important genes in Ashbya gossypii, we constructed Ashbya chromosome-specific BAC libraries using pBeloBAC11 and pBACwich vectors with an average insert size of 100 kb, which is equivalent to 19.8X genomic coverage. pBACwich was developed to streamline map-based cloning by providing a tool to integrate large DNA fragments into specific sites in chromosomes. These chromosome-specific libraries have provided a useful tool for the further characterization of the Ashbya genome including positional cloning and genome sequencing.
Ascomycota* ; Chromosomes, Artificial, Bacterial ; Chromosomes, Artificial, Yeast ; Clone Cells ; Cloning, Organism ; DNA ; Escherichia coli ; Gene Library ; Genome ; Mass Screening ; Plasmids

Cells, Cultured ; Hepatocytes ; transplantation ; Humans ; Liver ; cytology ; Liver, Artificial ; Stem Cells ; cytology

Fabricating human organs with tissue engineering and stem cells may be an alternative to current suboptimal therapies for treatment of malfunction or loss of human tissues or organs. From the tissue engineering's perspective, the patients are expected to be treated with new tissues or organs reconstructed with transplanted cells. The cells for tissue engineering could be somatic cells derived from the patients themselves, other individuals, or animals. Another valuable cell source would be stem cells. Embryonic stem cells retain the pluripotency to differentiate into every cell type of human organs, but ethical issues remain to be addressed. Adult stem cells may solve these ethical issues and immune rejection, but have limitation in differentiation into all ranges of tissue-specific cell types. Tissue engineering typically employs scaffolds fabricated from synthetic or natural biomaterials to engineer a new functional tissue from cells. The configuration of the biomaterials guides the structure of a regenerated tissue by defining a three-dimensional space. Appropriate combination of tissue engineering with stem cells shows a promise to fabricate human organs or tissues that can be utilized for patients in the near future.
Adult Stem Cells ; Animals ; Artificial Organs ; Biocompatible Materials ; Embryonic Stem Cells ; Ethics ; Humans ; Stem Cells* ; Tissue Engineering*

PURPOSE: The aim of this study was to evaluate the improvement of osteogenic potential of biphasic calcium phosphate (BCP) bone substitute coated with synthetic cell-binding peptide sequences in a standardized rabbit sinus model. METHODS: Standardized 6-mm diameter defects were created bilaterally on the maxillary sinus of ten male New Zealand white rabbits, receiving BCP bone substitute coated with synthetic cell binding peptide sequences on one side (experimental group) and BCP bone substitute without coating (control group) on the other side. Histologic and histomorphometric analysis of bone formation was carried out after a healing period of 4 or 8 weeks. RESULTS: Histological analysis revealed signs of new bone formation in both experimental groups (4- and 8-week healing groups) with a statistically significant increase in bone formation in the 4-week healing group compared to the control group. However, no statistically significant difference in bone formation was found between the 8-week healing group and the control group. CONCLUSIONS: This study found that BCP bone substitute coated with synthetic cell-binding peptide sequences enhanced osteoinductive potential in a standardized rabbit sinus model and its effectiveness was greater in the 4-week healing group than in the 8-week healing group.
Artificial Cells ; Bone Regeneration ; Bone Substitutes ; Calcium ; Durapatite ; Humans ; Hydroxyapatites ; Male ; Maxillary Sinus ; Oligopeptides ; Osteogenesis ; Rabbits

OBJECTIVETo investigate the feasibility of construction of tissue-engineered skin in vitro.

METHODSFibroblasts were seeded on the dermal surface of acellular dermal matrix (ADM). 7 days later, epidermal cells (5 x 10(5)/cm2) were mixed with fibroblasts (0.2 x 10(5)/cm2) and then seeded on epidermal surface of ADM. The culture medium was the mixture liquor containing K-SFM in half and the culture supernatants of fibroblasts in half. In the control group, only epidermal cells (5 x 10(5)/cm2) were seeded and cultured with K-SFM. After composite skin was cultured for 1 week and 3 week, samples were harvested respectively for morphological study and to receive identification by immunohistochemistry.

RESULTSAfter 3-week culture, there were 3-4 continuous layers of cells in the epidermis in the experiment group. The epidermis was attached tightly to the dermis with trochanterellus. But in the control group, there were just 1-2 layers of cells in the epidermis which was not connected to the dermis. Strong positive dye of Laminin indicated that basement membrane was thoroughly formed in the experiment group. The same result was demonstrated by transmission electron microscope.

CONCLUSIONSThe mix-seeding of epidermal cells and few fibroblasts would promote the adhesion and proliferation of epidermal cells on ADM, and the formation of basement membrane.

Animals ; Cells, Cultured ; Coculture Techniques ; Epidermis ; cytology ; Fibroblasts ; cytology ; Skin, Artificial ; Swine ; Tissue Engineering ; Tissue Scaffolds

OBJECTIVETo explore the separation and culture method of adult hepatocytes.

METHODSThe isolated adult hepatocytes were cultivated by RPMI 1640 medium at 37 degrees C in vitro. The characteristics of the growing hepatocytes were observed. Their synthesis of urea was detected. The transformation efficiency and density's change of lidocaine were analyzed.

RESULTSHepatocytes were successful separated from adult liver. And they were cultivated in common condition and hollow fiber reactor. The functional capacity of hepatocytes was for lidocaine metabolism and urea excretion.

CONCLUSIONThe adult hepatocytes have been successful separated from liver. And they can be cultivated in common condition and hollow fiber reactor. And it could provide a great quantity and high activity of hepatocytes for bioartificial liver.

Animals ; Cell Culture Techniques ; Cells, Cultured ; Hepatocytes ; metabolism ; Humans ; Liver ; cytology ; Liver, Artificial ; trends ; Research

OBJECTIVETo use a new kind of fixing material, i.e. Sol-Gel organic-inorganic hybridized material to immobilize bacterium to detect Biochemical oxygen demand quickly.

METHODSThe biosensor was fabricated using a thin film in which Hansenula anomala was immobilized by sol-gel and an oxygen electrode. The optimum measurement for biochemical oxygen demand was at pH 7.0; 28 degrees C; response time 3 - 12 min. Pure organic compound, sewage and rate of recovery were detected with the biosensor.

RESULTSIt shows that the BOD biosensor can be used to detect many organic compounds such as amino acid, glucide. It is suitable to monitor sewage and industrial waste water which has low level alcohols and phenols. The microbial membrane can work 3 months and remain its 70% activity. It is measured that the rate of recovery of BOD is between 90% to 105% in sewage.

CONCLUSIONThe study confirmed the effectiveness and usefulness of BOD sensor, which is quick, convenient, low cost and reliable with little interference.

Bacteria ; Biosensing Techniques ; instrumentation ; Cells, Immobilized ; Gels ; Membranes, Artificial ; Nylons ; Oxygen ; analysis ; Sewage ; analysis ; microbiology

This article presents the design of a bioreactor using hollow fiber membrane and, isolated hepatocytes of suckling pigs, and the experimental study of its efficacy in vitro. Liver cells were harvested from suckling pigs with collagenase perfusion in situ, and parenchymal and non-parenchymal hepatocytes were cocultured in a hollow fiber module which was rotated sporadically. Bioartificial liver(BAL) was developed using this bioreactor,and the BAL was perfused with ascites of patients suffering from liver cirrhosis. The yield of viable hepatocytes was (6.29 +/- 0.37) x 10(8) cells, and cell viability was greater than 84%. Hepatocytes aggregated to multi-cells spheroids after being rotated every thirty minutes for three hours. The hepatocytes in the bioreactor could synthesize urea. Total billirubin was decreased, and AST was significantly increased in the group of bioreactor, as compared with that in the control group. Glucose decreased in the group of bioreactor,whereas there was no significant descent in the control group; and the difference between the two groups was significant. The above results demonstrate that this bioreactor is effective for decreasing total bilirubin and glucose.
Animals ; Animals, Newborn ; Bioreactors ; Cell Separation ; Cells, Cultured ; Hepatocytes ; cytology ; Liver, Artificial ; Swine