We have developed two novel cell co-culture system, without any on cell type combination limitation, utilizing a polymer surface which is temperature-sensitive with respect to its cell adhesion characteristics. One system involves a patterned co-culture of primary hepatocytes with endothelial cells utilizing patterned masked of the electron-beam cured, temperature-responsive polymer, poly (N-isopropylacrylamide) (PIPAAm) by masked electron beam irradiation. Hepatocytes were cultured to confluency at 37 degrees C on these surfaces. When the culture temperature was reduced below 32 degrees C, cells detached from the PIPAAm-grafted areas without any need for trypsin. Endothelial cells were then seeded onto the same surfaces at 37 degrees C. These subsequently seeded endothelial cells adhered only to the now-exposed PIPAAm-grafted domains and could be co-cultured with the hepatocytes initially seeded at 37 degrees C in well-ordered patterns. The other system involves a double layered co-culture obtained by overlaying endothelial cell sheets of the designed shape onto hepatocyte monolayers. The endothelial cells adhered and proliferated on the PIPAAm-grafted surface, as on polystyrene tissue culture dishes at 37 degrees C. By reducing the temperature, confluent monolayers of cells detached from the PIPAAm surfaces without trypsin. Because the recovered cells maintaed intact cell-cell junctions together with deposited extracellular matrix, the harvested endothelial cell sheets, with designed shapes, were transferable and readily adhered to hepatocyte monolayers. Stable double layered cell sheets could be co-cultivated. These two co-culture methods enabled long-term co-culture of primary hepatocytes with endothelial cells. Hepatocytes so co-cultured with endothelial cells maintained their differentiated functions, such as albumin synthesis for unexpectedly long periods. These novel two co-culture systems offer promising techniques for basic biologic researches upon intercellular communications, and for the clinical applications of tissue engineered constructs.
Acrylic Resins/chemistry* ; Animal ; Coculture ; Cytological Techniques* ; Endothelium/cytology* ; Human ; Surface Properties ; Temperature*

BACKGROUND: Hepatocytes are an attractive cell source in hepatic tissue engineering because they are the primary cells of the liver, maintaining liver homeostasis through their intrinsic function. Due to the increasing demand for liver donors, a wide range of methods are being studied to obtain functionally active hepatocytes. iPSCs are one of the alternative cell sources, which shows great promise as a tool for generating hepatocytes. METHODS: This study determined whether factors associated with iPSCs contributed to variation in hepatocyte-like cells derived from iPSCs. The factors of concern for the iPSCs included the culture system, the source of iPSCs, and cell seeding density for initiating the differentiation. RESULTS: Our results found iPSC-dependent variances among differentiated hepatocyte-like cells. The matrix used in culturing iPSCs significantly impacts cell morphologies, characteristics, and the expression of pluripotent genes, such as OCT4 and SOX2, varied in iPSCs derived from different sources. These characteristics, in turn, play a consequential role in determining the functional activity of the iPSC-derived hepatocyte-like cells. In addition, cell seeding density was observed to be an essential factor for the efficient generation of iPSC-derived hepatocyte-like cells, with 2- 4 * 10 cells/cm of seeding density resulting in good morphology and functionality. CONCLUSION This study provides the baseline of effective differentiation protocols for iPSC-derived hepatocyte-like cells with the appropriate conditions, including cell culture media, iPSC source, and the seeding density of iPSCs.

BACKGROUND: Hepatocytes are an attractive cell source in hepatic tissue engineering because they are the primary cells of the liver, maintaining liver homeostasis through their intrinsic function. Due to the increasing demand for liver donors, a wide range of methods are being studied to obtain functionally active hepatocytes. iPSCs are one of the alternative cell sources, which shows great promise as a tool for generating hepatocytes. METHODS: This study determined whether factors associated with iPSCs contributed to variation in hepatocyte-like cells derived from iPSCs. The factors of concern for the iPSCs included the culture system, the source of iPSCs, and cell seeding density for initiating the differentiation. RESULTS: Our results found iPSC-dependent variances among differentiated hepatocyte-like cells. The matrix used in culturing iPSCs significantly impacts cell morphologies, characteristics, and the expression of pluripotent genes, such as OCT4 and SOX2, varied in iPSCs derived from different sources. These characteristics, in turn, play a consequential role in determining the functional activity of the iPSC-derived hepatocyte-like cells. In addition, cell seeding density was observed to be an essential factor for the efficient generation of iPSC-derived hepatocyte-like cells, with 2- 4 * 10 cells/cm of seeding density resulting in good morphology and functionality. CONCLUSION This study provides the baseline of effective differentiation protocols for iPSC-derived hepatocyte-like cells with the appropriate conditions, including cell culture media, iPSC source, and the seeding density of iPSCs.

BACKGROUND: Hepatocytes are an attractive cell source in hepatic tissue engineering because they are the primary cells of the liver, maintaining liver homeostasis through their intrinsic function. Due to the increasing demand for liver donors, a wide range of methods are being studied to obtain functionally active hepatocytes. iPSCs are one of the alternative cell sources, which shows great promise as a tool for generating hepatocytes. METHODS: This study determined whether factors associated with iPSCs contributed to variation in hepatocyte-like cells derived from iPSCs. The factors of concern for the iPSCs included the culture system, the source of iPSCs, and cell seeding density for initiating the differentiation. RESULTS: Our results found iPSC-dependent variances among differentiated hepatocyte-like cells. The matrix used in culturing iPSCs significantly impacts cell morphologies, characteristics, and the expression of pluripotent genes, such as OCT4 and SOX2, varied in iPSCs derived from different sources. These characteristics, in turn, play a consequential role in determining the functional activity of the iPSC-derived hepatocyte-like cells. In addition, cell seeding density was observed to be an essential factor for the efficient generation of iPSC-derived hepatocyte-like cells, with 2- 4 * 10 cells/cm of seeding density resulting in good morphology and functionality. CONCLUSION This study provides the baseline of effective differentiation protocols for iPSC-derived hepatocyte-like cells with the appropriate conditions, including cell culture media, iPSC source, and the seeding density of iPSCs.

BACKGROUND: Hepatocytes are an attractive cell source in hepatic tissue engineering because they are the primary cells of the liver, maintaining liver homeostasis through their intrinsic function. Due to the increasing demand for liver donors, a wide range of methods are being studied to obtain functionally active hepatocytes. iPSCs are one of the alternative cell sources, which shows great promise as a tool for generating hepatocytes. METHODS: This study determined whether factors associated with iPSCs contributed to variation in hepatocyte-like cells derived from iPSCs. The factors of concern for the iPSCs included the culture system, the source of iPSCs, and cell seeding density for initiating the differentiation. RESULTS: Our results found iPSC-dependent variances among differentiated hepatocyte-like cells. The matrix used in culturing iPSCs significantly impacts cell morphologies, characteristics, and the expression of pluripotent genes, such as OCT4 and SOX2, varied in iPSCs derived from different sources. These characteristics, in turn, play a consequential role in determining the functional activity of the iPSC-derived hepatocyte-like cells. In addition, cell seeding density was observed to be an essential factor for the efficient generation of iPSC-derived hepatocyte-like cells, with 2- 4 * 10 cells/cm of seeding density resulting in good morphology and functionality. CONCLUSION This study provides the baseline of effective differentiation protocols for iPSC-derived hepatocyte-like cells with the appropriate conditions, including cell culture media, iPSC source, and the seeding density of iPSCs.

BACKGROUND: Hepatocytes are an attractive cell source in hepatic tissue engineering because they are the primary cells of the liver, maintaining liver homeostasis through their intrinsic function. Due to the increasing demand for liver donors, a wide range of methods are being studied to obtain functionally active hepatocytes. iPSCs are one of the alternative cell sources, which shows great promise as a tool for generating hepatocytes. METHODS: This study determined whether factors associated with iPSCs contributed to variation in hepatocyte-like cells derived from iPSCs. The factors of concern for the iPSCs included the culture system, the source of iPSCs, and cell seeding density for initiating the differentiation. RESULTS: Our results found iPSC-dependent variances among differentiated hepatocyte-like cells. The matrix used in culturing iPSCs significantly impacts cell morphologies, characteristics, and the expression of pluripotent genes, such as OCT4 and SOX2, varied in iPSCs derived from different sources. These characteristics, in turn, play a consequential role in determining the functional activity of the iPSC-derived hepatocyte-like cells. In addition, cell seeding density was observed to be an essential factor for the efficient generation of iPSC-derived hepatocyte-like cells, with 2- 4 * 10 cells/cm of seeding density resulting in good morphology and functionality. CONCLUSION This study provides the baseline of effective differentiation protocols for iPSC-derived hepatocyte-like cells with the appropriate conditions, including cell culture media, iPSC source, and the seeding density of iPSCs.

Objective: To examine the trajectory of activities of daily living (ADL) in cancer patients from 6 weeks before death using the Functional Independence Measure (FIM). Method: This study was a retrospective observational study. The participants were cancer patients aged 18 years or older who died and were discharged from the palliative care unit in Tsurumaki-onsen Hospital. Six weeks of FIM data were collected from the patients’ medical records from 6 weeks before death to the time immediately before death (week 0). Results: Fifty-five participants were included in the study. FIM scores declined from 55 points at 6 weeks before death to 25 points immediately before death. Functional independence was higher for cognitive items on the FIM than for motor items, and both cognitive and motor functioning significantly declined just before death. Within the motor subscale, the patients were more independent with regard to eating, grooming, and bladder management compared to other activities until just before death. Within the cognitive subscale, the patients showed greater independence with regard to expression and social interaction. Discussion: A support for ADL needs to be considered to assist out-of bed activities in a safe and comfort manner until two weeks before death. Patients’ activities on the bed can also be continued for independence until just before death.