Pub Date : 2016-08-12DOI: 10.3727/215517916X692843
M. Lecomte, Séverine Pechberty, C. Machado, Sandra Da Barroca, P. Ravassard, R. Scharfmann, P. Czernichow, B. Duvillié
Innovative treatments to cure type 1 diabetes are being actively researched. Among the different strategies, the replacement of β-cells has given promising results. Classically, islets from cadaveric donors are transplanted into diabetic patients, but recently phase I clinical trials that use stem cell-derived β-cells have been started. Such protocols require either an immunosuppressive treatment or the macroencapsulation of the β-cells. They involve cell aggregation and the exposure of the cells to hypoxia. Using an engineered human β-cell, we have addressed these two problems: a novel human β-cell line called EndoC-βH3 was cultured as single cells or aggregated clusters. EndoC-βH3 cells were also cultured at normal atmospheric oxygen tension (pO2 = 21%) or hypoxia (pO2 = 3%) in the presence or absence of modulators of the hypoxia-inducible factor 1α (HIF1α) pathway. Cell aggregation improved glucose-stimulated insulin secretion, demonstrating the benefit of cell-cell contacts. Low oxygen tension decreased β-cell viability and their sensitivity to glucose, but did not alter insulin production nor the insulin secretion capacity of the remaining cells. To investigate the role of HIF1α, we first used a HIF stabilizer at pO2 = 21%. This led to a mild decrease in cell viability, impaired glucose sensitivity, and altered insulin secretion. Finally, we used a HIF inhibitor on EndoC-βH3 pseudoislets exposed to hypoxia. Such treatment considerably decreased cell viability. In conclusion, aggregation of the EndoC-βH3 cells seems to be important to improve their function. A fraction of the EndoC-βH3 cells are resistant to hypoxia, depending on the level of activity of HIF1α. Thus, these cells represent a good human cell model for future investigations on islet cell transplantation analysis.
{"title":"Aggregation of Engineered Human β-Cells Into Pseudoislets: Insulin Secretion and Gene Expression Profile in Normoxic and Hypoxic Milieu.","authors":"M. Lecomte, Séverine Pechberty, C. Machado, Sandra Da Barroca, P. Ravassard, R. Scharfmann, P. Czernichow, B. Duvillié","doi":"10.3727/215517916X692843","DOIUrl":"https://doi.org/10.3727/215517916X692843","url":null,"abstract":"Innovative treatments to cure type 1 diabetes are being actively researched. Among the different strategies, the replacement of β-cells has given promising results. Classically, islets from cadaveric donors are transplanted into diabetic patients, but recently phase I clinical trials that use stem cell-derived β-cells have been started. Such protocols require either an immunosuppressive treatment or the macroencapsulation of the β-cells. They involve cell aggregation and the exposure of the cells to hypoxia. Using an engineered human β-cell, we have addressed these two problems: a novel human β-cell line called EndoC-βH3 was cultured as single cells or aggregated clusters. EndoC-βH3 cells were also cultured at normal atmospheric oxygen tension (pO2 = 21%) or hypoxia (pO2 = 3%) in the presence or absence of modulators of the hypoxia-inducible factor 1α (HIF1α) pathway. Cell aggregation improved glucose-stimulated insulin secretion, demonstrating the benefit of cell-cell contacts. Low oxygen tension decreased β-cell viability and their sensitivity to glucose, but did not alter insulin production nor the insulin secretion capacity of the remaining cells. To investigate the role of HIF1α, we first used a HIF stabilizer at pO2 = 21%. This led to a mild decrease in cell viability, impaired glucose sensitivity, and altered insulin secretion. Finally, we used a HIF inhibitor on EndoC-βH3 pseudoislets exposed to hypoxia. Such treatment considerably decreased cell viability. In conclusion, aggregation of the EndoC-βH3 cells seems to be important to improve their function. A fraction of the EndoC-βH3 cells are resistant to hypoxia, depending on the level of activity of HIF1α. Thus, these cells represent a good human cell model for future investigations on islet cell transplantation analysis.","PeriodicalId":9780,"journal":{"name":"Cell medicine","volume":"8 3 1","pages":"99-112"},"PeriodicalIF":0.0,"publicationDate":"2016-08-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3727/215517916X692843","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"69756651","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2016-01-14DOI: 10.3727/215517916X690486
A. Grognuz, C. Scaletta, A. Farron, D. Pioletti, W. Raffoul, L. Applegate
Tendon afflictions are very common, and their negative impact is high both at the workplace and in leisure activities. Tendinopathies are increasing in prevalence and can lead to tendon ruptures, where healing is a long process with outcomes that are often disappointing. Human fetal progenitor tenocytes (hFPTs) have been recently tested in vitro as a potential cell source to stimulate tendon regeneration. The aim of the present study was to compare different commercial hyaluronic acid (HA) gels, which could be used to resuspend hFPTs in a formulation that would allow for good delivery of the cells. No medium or growth supplement was used in the formulation in order to make it therapeutically dispensable. These conditions are stringent for cells, but surprisingly, we found that different formulations could allow a good survival for up to 3 days when stored at 4°C (refrigerator stable). The gels must allow a good survival of the cells in parallel with a good stability of the preparation over time and sufficient viscosity to remain in place if deposited on a wounded location. Moreover, the cells must conserve their ability to attach and to proliferate. hFPTs were able to survive and to recover from all of the tested gels, but some products showed some advantages over others in terms of survival and viscosity. Finally, the Ostenil Tendon HA gel fulfilled all of the requirements and presented the best compromise between a good survival and sufficient rheological characteristics to create an interesting cell delivery system.
{"title":"Stability Enhancement Using Hyaluronic Acid Gels for Delivery of Human Fetal Progenitor Tenocytes.","authors":"A. Grognuz, C. Scaletta, A. Farron, D. Pioletti, W. Raffoul, L. Applegate","doi":"10.3727/215517916X690486","DOIUrl":"https://doi.org/10.3727/215517916X690486","url":null,"abstract":"Tendon afflictions are very common, and their negative impact is high both at the workplace and in leisure activities. Tendinopathies are increasing in prevalence and can lead to tendon ruptures, where healing is a long process with outcomes that are often disappointing. Human fetal progenitor tenocytes (hFPTs) have been recently tested in vitro as a potential cell source to stimulate tendon regeneration. The aim of the present study was to compare different commercial hyaluronic acid (HA) gels, which could be used to resuspend hFPTs in a formulation that would allow for good delivery of the cells. No medium or growth supplement was used in the formulation in order to make it therapeutically dispensable. These conditions are stringent for cells, but surprisingly, we found that different formulations could allow a good survival for up to 3 days when stored at 4°C (refrigerator stable). The gels must allow a good survival of the cells in parallel with a good stability of the preparation over time and sufficient viscosity to remain in place if deposited on a wounded location. Moreover, the cells must conserve their ability to attach and to proliferate. hFPTs were able to survive and to recover from all of the tested gels, but some products showed some advantages over others in terms of survival and viscosity. Finally, the Ostenil Tendon HA gel fulfilled all of the requirements and presented the best compromise between a good survival and sufficient rheological characteristics to create an interesting cell delivery system.","PeriodicalId":9780,"journal":{"name":"Cell medicine","volume":"8 3 1","pages":"87-97"},"PeriodicalIF":0.0,"publicationDate":"2016-01-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3727/215517916X690486","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"69756584","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-12-17DOI: 10.3727/215517915X689029
Shingo Yamashita, K. Ohashi, R. Utoh, T. Okano, Masakazu Yamamoto
Our experimental approach toward the development of new islet-based treatment for diabetes mellitus has been the creation of a monolayered islet cell construct (islet cell sheet), followed by its transplantation into a subcutaneous pocket. Previous studies describe rat laminin-5 (chain composition: α3, β3, γ2) as a suitable extracellular matrix (ECM) for surfaces comprised of a coated temperature-responsive polymer, poly(N-isopropylacrylamide) (PIPAAm). To progress toward the clinical application of this approach, the present study attempted to identify an optimal human ECM as a coating material on PIPAAm surfaces, which allowed islet cells to attach on the surfaces and subsequently to be harvested as a monolithic cell sheet. Dispersed rat islet cells were seeded onto PIPAAm dishes coated with various human laminin isotypes: human laminin (HL)-211, HL-332, HL-411, HL-511, and HL-placenta. Plating efficiency at day 1, the confluency at day 3, and glucose-stimulated insulin secretion test at day 3 were performed. The highest value of plating efficiency was found in the HL-332-PIPAAm group (83.1 ± 0.7%). The HL-332-PIPAAm group also showed the highest cellular confluency (98.6 ± 0.5%). Islet cells cultured on the HL-332-PIPAAm surfaces showed a positive response in the glucose-stimulated insulin secretion test. By reducing culture temperature from 37°C to 20°C in the HL-332-PIPAAm group, cells were able to be harvested as a monolithic islet sheet. The present study showed that HL-332 was an optimal human-derived ECM on a PIPAAm coating for preparing islet cell sheets.
{"title":"Human Laminin Isotype Coating for Creating Islet Cell Sheets.","authors":"Shingo Yamashita, K. Ohashi, R. Utoh, T. Okano, Masakazu Yamamoto","doi":"10.3727/215517915X689029","DOIUrl":"https://doi.org/10.3727/215517915X689029","url":null,"abstract":"Our experimental approach toward the development of new islet-based treatment for diabetes mellitus has been the creation of a monolayered islet cell construct (islet cell sheet), followed by its transplantation into a subcutaneous pocket. Previous studies describe rat laminin-5 (chain composition: α3, β3, γ2) as a suitable extracellular matrix (ECM) for surfaces comprised of a coated temperature-responsive polymer, poly(N-isopropylacrylamide) (PIPAAm). To progress toward the clinical application of this approach, the present study attempted to identify an optimal human ECM as a coating material on PIPAAm surfaces, which allowed islet cells to attach on the surfaces and subsequently to be harvested as a monolithic cell sheet. Dispersed rat islet cells were seeded onto PIPAAm dishes coated with various human laminin isotypes: human laminin (HL)-211, HL-332, HL-411, HL-511, and HL-placenta. Plating efficiency at day 1, the confluency at day 3, and glucose-stimulated insulin secretion test at day 3 were performed. The highest value of plating efficiency was found in the HL-332-PIPAAm group (83.1 ± 0.7%). The HL-332-PIPAAm group also showed the highest cellular confluency (98.6 ± 0.5%). Islet cells cultured on the HL-332-PIPAAm surfaces showed a positive response in the glucose-stimulated insulin secretion test. By reducing culture temperature from 37°C to 20°C in the HL-332-PIPAAm group, cells were able to be harvested as a monolithic islet sheet. The present study showed that HL-332 was an optimal human-derived ECM on a PIPAAm coating for preparing islet cell sheets.","PeriodicalId":9780,"journal":{"name":"Cell medicine","volume":"8 1-2 1","pages":"39-46"},"PeriodicalIF":0.0,"publicationDate":"2015-12-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3727/215517915X689029","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"69756403","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-12-17DOI: 10.3727/215517915X689100
Chika Miyagi-Shiohira, Kiyoto Kurima, N. Kobayashi, I. Saitoh, Masami Watanabe, Yasufumi Noguchi, Masayuki Matsushita, H. Noguchi
Mesenchymal stem cells (MSCs) have the potential to differentiate into cells of mesodermal origin such as osteoblasts, adipocytes, myocytes, and chondrocytes. They possess an immunosuppressive effect, which makes them a viable cell population for the cell-based therapy of treatment-resistant immune diseases. Adipose-derived mesenchymal stem cells (ASCs) have been demonstrated to have the ability to acquire the properties of subcutaneous adipose tissue particularly easily, and cryopreservation is currently performed as a routine method for preserving ASCs to safely acquire large numbers of cells. However, many studies have reported that cellular activity after freezing and thawing may be affected by the solutions used for cryopreservation. Dimethyl sulfoxide (DMSO) is commonly used as a cryopreservation medium as it diffuses into the cell through the plasma membrane and protects the cells from the damage caused by freezing. As substitutes for DMSO or animal-derived serum, cell banker series, polyvinylpyrrolidone (PVP), sericin and maltose, and methyl cellulose (MC) have been investigated for their clinical applications. It is critical to develop a reliable cell cryopreservation protocol for regenerative medicine using MSCs.
{"title":"Cryopreservation of Adipose-Derived Mesenchymal Stem Cells.","authors":"Chika Miyagi-Shiohira, Kiyoto Kurima, N. Kobayashi, I. Saitoh, Masami Watanabe, Yasufumi Noguchi, Masayuki Matsushita, H. Noguchi","doi":"10.3727/215517915X689100","DOIUrl":"https://doi.org/10.3727/215517915X689100","url":null,"abstract":"Mesenchymal stem cells (MSCs) have the potential to differentiate into cells of mesodermal origin such as osteoblasts, adipocytes, myocytes, and chondrocytes. They possess an immunosuppressive effect, which makes them a viable cell population for the cell-based therapy of treatment-resistant immune diseases. Adipose-derived mesenchymal stem cells (ASCs) have been demonstrated to have the ability to acquire the properties of subcutaneous adipose tissue particularly easily, and cryopreservation is currently performed as a routine method for preserving ASCs to safely acquire large numbers of cells. However, many studies have reported that cellular activity after freezing and thawing may be affected by the solutions used for cryopreservation. Dimethyl sulfoxide (DMSO) is commonly used as a cryopreservation medium as it diffuses into the cell through the plasma membrane and protects the cells from the damage caused by freezing. As substitutes for DMSO or animal-derived serum, cell banker series, polyvinylpyrrolidone (PVP), sericin and maltose, and methyl cellulose (MC) have been investigated for their clinical applications. It is critical to develop a reliable cell cryopreservation protocol for regenerative medicine using MSCs.","PeriodicalId":9780,"journal":{"name":"Cell medicine","volume":"8 1-2 1","pages":"3-7"},"PeriodicalIF":0.0,"publicationDate":"2015-12-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3727/215517915X689100","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"69756576","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-12-17DOI: 10.3727/215517915X689038
I. Saitoh, E. Inada, Y. Iwase, H. Noguchi, Tomoya Murakami, Miki Soda, Naoko Kubota, H. Hasegawa, Eri Akasaka, Y. Matsumoto, K. Oka, Y. Yamasaki, H. Hayasaki, Masahiro Sato
Feeder cells are generally required to maintain embryonic stem cells (ESCs)/induced pluripotent stem cells (iPSCs). Mouse embryonic fibroblasts (MEFs) isolated from fetuses and STO mouse stromal cell line are the most widely used feeder cells. The aim of this study was to determine which cells are suitable for establishing iPSCs from human deciduous tooth dental pulp cells (HDDPCs). Primary cultures of HDDPCs were cotransfected with three plasmids containing human OCT3/4, SOX2/KLF4, or LMYC/LIN28 and pmaxGFP by using a novel electroporation method, and then cultured in an ESC qualified medium for 15 days. Emerging colonies were reseeded onto mitomycin C-treated MEFs or STO cells. The colonies were serially passaged for up to 26 passages. During this period, colony morphology was assessed to determine whether cells exhibited ESC-like morphology and alkaline phosphatase activity to evaluate the state of cellular reprogramming. HDDPCs maintained on MEFs were successfully reprogrammed into iPSCs, whereas those maintained on STO cells were not. Once established, the iPSCs were maintained on STO cells without loss of pluripotency. Our results indicate that MEFs are better feeder cells than STO cells for establishing iPSCs. Feeder choice is a key factor enabling efficient generation of iPSCs.
{"title":"Choice of Feeders Is Important When First Establishing iPSCs Derived From Primarily Cultured Human Deciduous Tooth Dental Pulp Cells.","authors":"I. Saitoh, E. Inada, Y. Iwase, H. Noguchi, Tomoya Murakami, Miki Soda, Naoko Kubota, H. Hasegawa, Eri Akasaka, Y. Matsumoto, K. Oka, Y. Yamasaki, H. Hayasaki, Masahiro Sato","doi":"10.3727/215517915X689038","DOIUrl":"https://doi.org/10.3727/215517915X689038","url":null,"abstract":"Feeder cells are generally required to maintain embryonic stem cells (ESCs)/induced pluripotent stem cells (iPSCs). Mouse embryonic fibroblasts (MEFs) isolated from fetuses and STO mouse stromal cell line are the most widely used feeder cells. The aim of this study was to determine which cells are suitable for establishing iPSCs from human deciduous tooth dental pulp cells (HDDPCs). Primary cultures of HDDPCs were cotransfected with three plasmids containing human OCT3/4, SOX2/KLF4, or LMYC/LIN28 and pmaxGFP by using a novel electroporation method, and then cultured in an ESC qualified medium for 15 days. Emerging colonies were reseeded onto mitomycin C-treated MEFs or STO cells. The colonies were serially passaged for up to 26 passages. During this period, colony morphology was assessed to determine whether cells exhibited ESC-like morphology and alkaline phosphatase activity to evaluate the state of cellular reprogramming. HDDPCs maintained on MEFs were successfully reprogrammed into iPSCs, whereas those maintained on STO cells were not. Once established, the iPSCs were maintained on STO cells without loss of pluripotency. Our results indicate that MEFs are better feeder cells than STO cells for establishing iPSCs. Feeder choice is a key factor enabling efficient generation of iPSCs.","PeriodicalId":9780,"journal":{"name":"Cell medicine","volume":"8 1-2 1","pages":"9-23"},"PeriodicalIF":0.0,"publicationDate":"2015-12-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3727/215517915X689038","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"69756463","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-10-01DOI: 10.3727/215517915X688606
D. Eve
A wide variety of subjects are presented at the annual American Society of Neural Therapy and Repair meeting every year, as typified by this summary of the 2014 meeting. Parkinson's disease-related presentations were again the most popular topic, with traumatic brain injury, spinal cord injury, and stroke being close behind. Other disorders included Huntington's disease, brain cancer, and bipolar disorders. Several studies were related to multiple diseases, and many studies attempted to reveal more about the disease process. The use of scaffolds, drugs, and gene therapy as disease models and/or potential therapies were also featured. An increasing proportion of presentations related to stem cells, with the study of multiple stem cell types being the most common. Induced pluripotent stem cells were increasingly popular, including two presentations each on a muscle-derived dedifferentiated cell type and cells derived from bipolar patients. Other stem cells, including neural stem cells, mesenchymal stem cells, umbilical cord blood cells, and embryonic stem cells, were featured. More than 55% of the stem cell studies involved transplantation, with human-derived cells being the most frequently transplanted, while rats were the most common recipient. Two human autologous studies for spinal cord injury and hypoxia-derived encephalopathy, while a further three allogenic studies for stroke and spinal cord injury, were also featured. This year's meeting highlights the increasing promise of stem cells and other therapies for the treatment of neurodegenerative disorders.
{"title":"Disease and Stem Cell-Based Analysis of the 2014 ASNTR Meeting.","authors":"D. Eve","doi":"10.3727/215517915X688606","DOIUrl":"https://doi.org/10.3727/215517915X688606","url":null,"abstract":"A wide variety of subjects are presented at the annual American Society of Neural Therapy and Repair meeting every year, as typified by this summary of the 2014 meeting. Parkinson's disease-related presentations were again the most popular topic, with traumatic brain injury, spinal cord injury, and stroke being close behind. Other disorders included Huntington's disease, brain cancer, and bipolar disorders. Several studies were related to multiple diseases, and many studies attempted to reveal more about the disease process. The use of scaffolds, drugs, and gene therapy as disease models and/or potential therapies were also featured. An increasing proportion of presentations related to stem cells, with the study of multiple stem cell types being the most common. Induced pluripotent stem cells were increasingly popular, including two presentations each on a muscle-derived dedifferentiated cell type and cells derived from bipolar patients. Other stem cells, including neural stem cells, mesenchymal stem cells, umbilical cord blood cells, and embryonic stem cells, were featured. More than 55% of the stem cell studies involved transplantation, with human-derived cells being the most frequently transplanted, while rats were the most common recipient. Two human autologous studies for spinal cord injury and hypoxia-derived encephalopathy, while a further three allogenic studies for stroke and spinal cord injury, were also featured. This year's meeting highlights the increasing promise of stem cells and other therapies for the treatment of neurodegenerative disorders.","PeriodicalId":9780,"journal":{"name":"Cell medicine","volume":"35 1","pages":"133-42"},"PeriodicalIF":0.0,"publicationDate":"2015-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3727/215517915X688606","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"69756324","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-10-01DOI: 10.3727/215517914X681794
M. Raredon, M. Ghaedi, Elizabeth A. Calle, L. Niklason
Respiratory epithelium is difficult to grow in vitro, as it requires a well-maintained polarizing air-liquid interface (ALI) to maintain differentiation. Traditional methods rely on permeable membrane culture inserts, which are difficult to work with and are ill-suited for the production of large numbers of cells, such as the quantities required for cell-based clinical therapies. Herein, we investigate an alternative form of culture in which the cells are placed on a porous substrate that is continuously rolled, such that the monolayer of cells is alternately submerged in media or apically exposed to air. Our prototype bioreactor is reliable for up to 21 days of continuous culture and is designed for scale-up for large-scale cell culture with continuous medium and gas exchange. Normal human bronchial epithelial (NHBE) cells were cultured on an absorbent substrate in the reactor for periods of 7, 14, and 21 days and were compared to static controls that were submerged in media. Quantification by immunohistochemistry and quantitative PCR of markers specific to differentiated respiratory epithelium indicated increased cilia, mucous production, and tight junction formation in the rolled cultures, compared to static. Together with scanning electron microscopy and paraffin histology, the data indicate that the intermittent ALI provided by the rolling bioreactor promotes a polarized epithelial phenotype over a period of 21 days.
{"title":"A Rotating Bioreactor for Scalable Culture and Differentiation of Respiratory Epithelium.","authors":"M. Raredon, M. Ghaedi, Elizabeth A. Calle, L. Niklason","doi":"10.3727/215517914X681794","DOIUrl":"https://doi.org/10.3727/215517914X681794","url":null,"abstract":"Respiratory epithelium is difficult to grow in vitro, as it requires a well-maintained polarizing air-liquid interface (ALI) to maintain differentiation. Traditional methods rely on permeable membrane culture inserts, which are difficult to work with and are ill-suited for the production of large numbers of cells, such as the quantities required for cell-based clinical therapies. Herein, we investigate an alternative form of culture in which the cells are placed on a porous substrate that is continuously rolled, such that the monolayer of cells is alternately submerged in media or apically exposed to air. Our prototype bioreactor is reliable for up to 21 days of continuous culture and is designed for scale-up for large-scale cell culture with continuous medium and gas exchange. Normal human bronchial epithelial (NHBE) cells were cultured on an absorbent substrate in the reactor for periods of 7, 14, and 21 days and were compared to static controls that were submerged in media. Quantification by immunohistochemistry and quantitative PCR of markers specific to differentiated respiratory epithelium indicated increased cilia, mucous production, and tight junction formation in the rolled cultures, compared to static. Together with scanning electron microscopy and paraffin histology, the data indicate that the intermittent ALI provided by the rolling bioreactor promotes a polarized epithelial phenotype over a period of 21 days.","PeriodicalId":9780,"journal":{"name":"Cell medicine","volume":"7 3 1","pages":"109-21"},"PeriodicalIF":0.0,"publicationDate":"2015-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3727/215517914X681794","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"69756558","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-10-01DOI: 10.3727/215517914X680038
M. A. Barros, J. P. F. Martins, D. Maria, Crisitiane Valverde Wenceslau, Dener Madeiro de Souza, A. Kerkis, N. Câmara, J. Balieiro, I. Kerkis
Acute renal failure (ARF) is a common renal disease that can lead to high mortality. Recovery from ARF occurs with the replacement of necrotic tubular cells by functional tubular epithelial cells and the normalization of microvascular endothelial cell function in the peritubular capillaries. Conventional therapeutic techniques are often ineffective against ARF. Hence, stem cell therapies, which act through multiple trophic and regenerative mechanisms, are encouraging. We investigated the homing of human immature dental pulp stem cells (IDPSCs) after endovenous (EV) or intraperitoneal (IP) injection, in immunocompetent Wistar rats with ARF induced by intramuscular injection of glycerol, without the use of immunosuppression. The cells, which had been cryopreserved for 6 years, were CD105(+), CD73(+), CD44(+), and partly, STRO-1(+) and CD146(+), and presented unaltered mesoderm differentiation potential. The presence of these cells in the tubular region of the kidney and in the peritubular capillaries was demonstrated. These cells accelerate tubular epithelial cell regeneration through significant increase of Ki-67-immunoreactive cells in damaged kidney. Flow cytometry analysis confirmed that IDPSCs home to the kidneys (EV 34.10% and IP 33.25%); a lower percentage of cells was found in the liver (EV 19.05% and IP 9.10%), in the muscles (EV 6.30% and IP 1.35%), and in the lungs (EV 2.0% and IP 1.85%). After infusion into rat, these cells express pericyte markers, such as CD146(+), STRO-1(+), and vascular endothelial growth factor (VEGF(+)). We found that IDPSCs demonstrate renotropic and pericyte-like properties and contributed to restore renal tubule structure in an experimental rat ARF model.
{"title":"Immature Dental Pulp Stem Cells Showed Renotropic and Pericyte-Like Properties in Acute Renal Failure in Rats.","authors":"M. A. Barros, J. P. F. Martins, D. Maria, Crisitiane Valverde Wenceslau, Dener Madeiro de Souza, A. Kerkis, N. Câmara, J. Balieiro, I. Kerkis","doi":"10.3727/215517914X680038","DOIUrl":"https://doi.org/10.3727/215517914X680038","url":null,"abstract":"Acute renal failure (ARF) is a common renal disease that can lead to high mortality. Recovery from ARF occurs with the replacement of necrotic tubular cells by functional tubular epithelial cells and the normalization of microvascular endothelial cell function in the peritubular capillaries. Conventional therapeutic techniques are often ineffective against ARF. Hence, stem cell therapies, which act through multiple trophic and regenerative mechanisms, are encouraging. We investigated the homing of human immature dental pulp stem cells (IDPSCs) after endovenous (EV) or intraperitoneal (IP) injection, in immunocompetent Wistar rats with ARF induced by intramuscular injection of glycerol, without the use of immunosuppression. The cells, which had been cryopreserved for 6 years, were CD105(+), CD73(+), CD44(+), and partly, STRO-1(+) and CD146(+), and presented unaltered mesoderm differentiation potential. The presence of these cells in the tubular region of the kidney and in the peritubular capillaries was demonstrated. These cells accelerate tubular epithelial cell regeneration through significant increase of Ki-67-immunoreactive cells in damaged kidney. Flow cytometry analysis confirmed that IDPSCs home to the kidneys (EV 34.10% and IP 33.25%); a lower percentage of cells was found in the liver (EV 19.05% and IP 9.10%), in the muscles (EV 6.30% and IP 1.35%), and in the lungs (EV 2.0% and IP 1.85%). After infusion into rat, these cells express pericyte markers, such as CD146(+), STRO-1(+), and vascular endothelial growth factor (VEGF(+)). We found that IDPSCs demonstrate renotropic and pericyte-like properties and contributed to restore renal tubule structure in an experimental rat ARF model.","PeriodicalId":9780,"journal":{"name":"Cell medicine","volume":"7 3 1","pages":"95-108"},"PeriodicalIF":0.0,"publicationDate":"2015-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3727/215517914X680038","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"69756447","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-08-26DOI: 10.3727/215517915X689047
H. Noguchi, Chika Miyagi-Shiohira, Kiyoto Kurima, N. Kobayashi, I. Saitoh, Masami Watanabe, Yasufumi Noguchi, Masayuki Matsushita
Although islet culture prior to transplantation provides flexibility for the evaluation of isolated islets and the pretreatment of patients, it is well known that isolated islets deteriorate rapidly in culture. Human serum albumin (HSA) is used for medium supplementation instead of fetal bovine serum (FBS), which is typically used for islet culture research, to avoid the introduction of xenogeneic materials. However, FBS contains several factors that are beneficial to islet viability and which also neutralize the endogenous pancreatic enzymes or exogenous enzymes left over from the isolation process. Several groups have reported the comparison of cultures at 22°C and 37°C. Recent studies have demonstrated the superiority of 4°C preservation to 22°C and 37°C cultures. We herein review the current research on islet culture/preservation for clinical islet transplantation.
{"title":"Islet Culture/Preservation Before Islet Transplantation.","authors":"H. Noguchi, Chika Miyagi-Shiohira, Kiyoto Kurima, N. Kobayashi, I. Saitoh, Masami Watanabe, Yasufumi Noguchi, Masayuki Matsushita","doi":"10.3727/215517915X689047","DOIUrl":"https://doi.org/10.3727/215517915X689047","url":null,"abstract":"Although islet culture prior to transplantation provides flexibility for the evaluation of isolated islets and the pretreatment of patients, it is well known that isolated islets deteriorate rapidly in culture. Human serum albumin (HSA) is used for medium supplementation instead of fetal bovine serum (FBS), which is typically used for islet culture research, to avoid the introduction of xenogeneic materials. However, FBS contains several factors that are beneficial to islet viability and which also neutralize the endogenous pancreatic enzymes or exogenous enzymes left over from the isolation process. Several groups have reported the comparison of cultures at 22°C and 37°C. Recent studies have demonstrated the superiority of 4°C preservation to 22°C and 37°C cultures. We herein review the current research on islet culture/preservation for clinical islet transplantation.","PeriodicalId":9780,"journal":{"name":"Cell medicine","volume":"8 1-2 1","pages":"25-9"},"PeriodicalIF":0.0,"publicationDate":"2015-08-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3727/215517915X689047","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"69756473","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-08-26DOI: 10.3727/215517915X689010
H. Noguchi
On behalf of the Japan Society for Organ Preservation and Medical Biology (JSOPMB), I express my sincere appreciation to Dr. David J. Eve and Professor Cesar V. Borlongan, Coeditors of Cell Medicine, for providing us with such an excellent opportunity to publish the data that were presented at the annual meeting of the JSOPMB. I also thank Dr. David J. Eve for the editing of our articles in detail. I am very sure that the relationship between Cell Medicine and JSOPMB has enhanced the motivation of JSOPMB members as well as board members and will continue to do so in the future, while also encouraging young Japanese researchers to join this organization. One of the extremely important missions of the annual meeting of the JSOPMB is to exchange new research outcomes and create new therapeutic concepts. JSOPMB always encourages and motivates young investigators. JSOPMB was started in 1974 for the study of organ preservation and developed widely in the 1990s with the participation of researchers in various fields of medicine, pharmacology, engineering, veterinary medicine, and basic science. Currently, JSOPMB has more than 700 members and is run under the direction of Professor Takashi Kondo, the president of the JSOPMB. Excellent presentations conducted at the 41th annual meeting of the JSOPMB held on November 28–29, 2014, in Osaka, Japan, under the supervision of Dr. Yoshiki Sawa (Professor, Department of Cardiovascular Surgery, Osaka University Graduate School of Medicine, Osaka, Japan), were selected and given an opportunity to be published in this special issue of Cell Medicine. Seven of these presentations are herein published in this special JSOPMB issue. Stem cell research was a major topic of interest. There were two articles regarding stem cells. Miyagi-Shiohira et al. reviewed cryopreservation of mesenchymal stem cells, especially adipose-derived stem cells. Dimethyl sulfoxide (DMSO) is commonly used as a cryopreservation medium as it diffuses into the cell through the plasma membrane and protects the cells from the damage caused by freezing. As substitutes for DMSO or animal-derived serum, cell banker series, polyvinylpyrrolidone (PVP), sericin and maltose, and methyl cellulose (MC) have been investigated for their clinical applications. Saitoh et al. evaluated feeder cells for establishing induced pluripotent stem cells (iPSCs) from human deciduous tooth dental pulp cells (HDDPCs). Their data showed that mouse embryonic fibroblasts (MEFs) were better feeder cells than an immortalized line established from Santos inbred mouse (SIM) embryonic fibroblasts resistant to 6-thioguanine and ouabain known as STO cells for establishing iPSCs. There were three articles regarding pancreatic islets. Noguchi et al. reviewed islet culture/preservation before islet transplantation. In the current clinical culture conditions, it seems that islets cultured at 37°C cannot receive enough oxygen because central necrosis of islets during culture w
我谨代表日本器官保存与医学生物学学会(JSOPMB)向《细胞医学》的共同编辑David J. Eve博士和Cesar V. Borlongan教授表示衷心的感谢，感谢他们为我们提供了这样一个极好的机会来发表在JSOPMB年会上提交的数据。我还要感谢David J. Eve博士对我们文章的详细编辑。我非常确信，细胞医学与JSOPMB之间的关系增强了JSOPMB成员和董事会成员的动力，并将在未来继续这样做，同时也鼓励年轻的日本研究人员加入这个组织。JSOPMB年会的一个极其重要的任务是交流新的研究成果和创造新的治疗理念。JSOPMB一直鼓励和激励年轻的调查人员。JSOPMB成立于1974年，目的是研究器官保存，在20世纪90年代得到了医学、药理学、工程、兽医学和基础科学等各个领域的研究人员的广泛发展。目前，JSOPMB有700多名会员，由JSOPMB会长近藤隆教授领导。2014年11月28日至29日在日本大阪举行的JSOPMB第41届年会上，在Yoshiki Sawa博士(日本大阪大学医学院心血管外科教授)的监督下，优秀的报告被选中并有机会发表在这期《细胞医学》特刊上。其中7个演讲将在本期JSOPMB特刊中发表。干细胞研究是一个重要的话题。有两篇关于干细胞的文章。Miyagi-Shiohira等人回顾了间充质干细胞的冷冻保存，特别是脂肪来源的干细胞。二甲基亚砜(DMSO)通常被用作低温保存介质，因为它通过质膜扩散到细胞中，保护细胞免受冷冻造成的损害。作为DMSO或动物源性血清的替代品，细胞库系列、聚乙烯吡罗烷酮(PVP)、丝胶蛋白和麦芽糖以及甲基纤维素(MC)已被研究用于临床应用。saiitoh等人评估了从人乳牙牙髓细胞(HDDPCs)培养诱导多能干细胞(iPSCs)的饲养细胞。他们的数据表明，小鼠胚胎成纤维细胞(mef)比桑托斯近交小鼠(SIM)胚胎成纤维细胞(STO细胞)建立的永生系更好地饲养细胞，用于建立iPSCs。有三篇关于胰岛的文章。Noguchi等人回顾了胰岛移植前的胰岛培养/保存。在目前的临床培养条件下，37℃培养的胰岛似乎无法获得足够的氧气，因为在培养过程中经常观察到胰岛中枢坏死。低温储存可能比培养更好，尽管未来的研究应该仔细评估胰岛当量和组织体积对4°c保存的胰岛抗原性的影响。Hanayama等人利用纤维修饰的腺病毒载体，在纤维结节的c端携带聚赖氨酸(K7)肽，证明了在培养中分散的胰岛细胞的有效基因转导。纤维修饰的腺病毒载体在感染倍数(MOI)为5和10时比传统的腺病毒载体具有更高的转导效率。Yamashita等人评估了人层粘连蛋白(HL)同型涂层用于制造胰岛细胞片的效果。HL-332是一种最佳的人源性细胞外基质(ECM)，用于涂层温度敏感的聚合物，聚n-异丙基丙烯酰胺(PIPAAm)表面。Miyamoto等人展示了一种用于集群培养的锥形模板(TASCL)装置，该装置用于体外培养肝球体。TASCL装置可作为药物毒性评价系统。Pillai等人利用链亲和素- qds585和生物素-pep-BHQ-1将黑洞猝灭剂(BHQ)分子偶联，以肽为中介，实现了量子点(QDs)的荧光关闭状态。该技术有望用于QD荧光开关控制的分子成像。这期JSOPMB的主题是“创造移植的未来”。董事会成员和我都期待着JSOPMB与细胞医学联合取得进一步进展。
{"title":"Creating a Future of Transplantation.","authors":"H. Noguchi","doi":"10.3727/215517915X689010","DOIUrl":"https://doi.org/10.3727/215517915X689010","url":null,"abstract":"On behalf of the Japan Society for Organ Preservation and Medical Biology (JSOPMB), I express my sincere appreciation to Dr. David J. Eve and Professor Cesar V. Borlongan, Coeditors of Cell Medicine, for providing us with such an excellent opportunity to publish the data that were presented at the annual meeting of the JSOPMB. I also thank Dr. David J. Eve for the editing of our articles in detail. I am very sure that the relationship between Cell Medicine and JSOPMB has enhanced the motivation of JSOPMB members as well as board members and will continue to do so in the future, while also encouraging young Japanese researchers to join this organization. \u0000 \u0000One of the extremely important missions of the annual meeting of the JSOPMB is to exchange new research outcomes and create new therapeutic concepts. JSOPMB always encourages and motivates young investigators. JSOPMB was started in 1974 for the study of organ preservation and developed widely in the 1990s with the participation of researchers in various fields of medicine, pharmacology, engineering, veterinary medicine, and basic science. Currently, JSOPMB has more than 700 members and is run under the direction of Professor Takashi Kondo, the president of the JSOPMB. \u0000 \u0000Excellent presentations conducted at the 41th annual meeting of the JSOPMB held on November 28–29, 2014, in Osaka, Japan, under the supervision of Dr. Yoshiki Sawa (Professor, Department of Cardiovascular Surgery, Osaka University Graduate School of Medicine, Osaka, Japan), were selected and given an opportunity to be published in this special issue of Cell Medicine. Seven of these presentations are herein published in this special JSOPMB issue. \u0000 \u0000Stem cell research was a major topic of interest. There were two articles regarding stem cells. Miyagi-Shiohira et al. reviewed cryopreservation of mesenchymal stem cells, especially adipose-derived stem cells. Dimethyl sulfoxide (DMSO) is commonly used as a cryopreservation medium as it diffuses into the cell through the plasma membrane and protects the cells from the damage caused by freezing. As substitutes for DMSO or animal-derived serum, cell banker series, polyvinylpyrrolidone (PVP), sericin and maltose, and methyl cellulose (MC) have been investigated for their clinical applications. Saitoh et al. evaluated feeder cells for establishing induced pluripotent stem cells (iPSCs) from human deciduous tooth dental pulp cells (HDDPCs). Their data showed that mouse embryonic fibroblasts (MEFs) were better feeder cells than an immortalized line established from Santos inbred mouse (SIM) embryonic fibroblasts resistant to 6-thioguanine and ouabain known as STO cells for establishing iPSCs. \u0000 \u0000There were three articles regarding pancreatic islets. Noguchi et al. reviewed islet culture/preservation before islet transplantation. In the current clinical culture conditions, it seems that islets cultured at 37°C cannot receive enough oxygen because central necrosis of islets during culture w","PeriodicalId":9780,"journal":{"name":"Cell medicine","volume":"8 1-2 1","pages":"1"},"PeriodicalIF":0.0,"publicationDate":"2015-08-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3727/215517915X689010","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"69756392","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
京公网安备 11010802042870号
京ICP备2023020795号-1
扫码关注我们
求助内容：
应助结果提醒方式：