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Derrick E. Rancourt - One of the best experts on this subject based on the ideXlab platform.
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An Integrated Approach toward the Biomanufacturing of Engineered Cell Therapy Products in a Stirred-Suspension Bioreactor.
Molecular therapy. Methods & clinical development, 2018Co-Authors: Charlie Y.m. Hsu, Tylor Walsh, Breanna S. Borys, Michael S. Kallos, Derrick E. RancourtAbstract:Recent advances in stem Cell biology have accelerated the pre-clinical development of Cell-based therapies for degenerative and chronic diseases. The success of this growing area hinges upon the concomitant development of scalable manufacturing platforms that can produce clinically relevant quantities of Cells for thousands of patients. Current biomanufacturing practices for Cell therapy products are built on a model previously optimized for biologics, wherein stable Cell lines are established first, followed by large-scale production in the bioreactor. This "two-step" approach can be costly, labor-intensive, and time-consuming, particularly for Cell therapy products that must be individually sourced from patients or compatible donors. In this report, we describe a "one-step" integrated approach toward the biomanufacturing of Engineered Cell therapy products by direct transfection of primary human fibroblast in a continuous stirred-suspension bioreactor. We optimized the transfection efficiency by testing rate-limiting factors, including Cell seeding density, agitation rate, oxygen saturation, microcarrier type, and serum concentration. By combining the genetic modification step with the large-scale expansion step, this not only removes the need for manual handing of Cells in planar culture dishes, but also enables the biomanufacturing process to be streamlined and automated in one fully enclosed bioreactor.
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An Integrated Approach toward the Biomanufacturing of Engineered Cell Therapy Products in a Stirred-Suspension Bioreactor
Elsevier, 2018Co-Authors: Charlie Y.m. Hsu, Tylor Walsh, Breanna S. Borys, Michael S. Kallos, Derrick E. RancourtAbstract:Recent advances in stem Cell biology have accelerated the pre-clinical development of Cell-based therapies for degenerative and chronic diseases. The success of this growing area hinges upon the concomitant development of scalable manufacturing platforms that can produce clinically relevant quantities of Cells for thousands of patients. Current biomanufacturing practices for Cell therapy products are built on a model previously optimized for biologics, wherein stable Cell lines are established first, followed by large-scale production in the bioreactor. This “two-step” approach can be costly, labor-intensive, and time-consuming, particularly for Cell therapy products that must be individually sourced from patients or compatible donors. In this report, we describe a “one-step” integrated approach toward the biomanufacturing of Engineered Cell therapy products by direct transfection of primary human fibroblast in a continuous stirred-suspension bioreactor. We optimized the transfection efficiency by testing rate-limiting factors, including Cell seeding density, agitation rate, oxygen saturation, microcarrier type, and serum concentration. By combining the genetic modification step with the large-scale expansion step, this not only removes the need for manual handing of Cells in planar culture dishes, but also enables the biomanufacturing process to be streamlined and automated in one fully enclosed bioreactor. Keywords: bioprocess, gene delivery, transfection, biotechnology, cationic polymer, Cell therapy, biomanufacturing, genetic engineering, plasmid DNA, bioreacto
Kohji Nishida - One of the best experts on this subject based on the ideXlab platform.
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Ebselen Preserves Tissue-Engineered Cell Sheets and their Stem Cells in Hypothermic Conditions.
Scientific reports, 2016Co-Authors: Ryosuke Katori, Ryuhei Hayashi, Yuki Kobayashi, Eiji Kobayashi, Kohji NishidaAbstract:Clinical trials have been performed using autologous tissue-Engineered epithelial Cell sheets for corneal regenerative medicine. To improve stem Cell-based therapy for convenient clinical practice, new techniques are required for preserving reconstructed tissues and their stem/progenitor Cells until they are ready for use. In the present study, we screened potential preservative agents and developed a novel medium for preserving the Cell sheets and their stem/progenitor Cells; the effects were evaluated with a luciferase-based viability assay. Nrf2 activators, specifically ebselen, could maintain high ATP levels during preservation. Ebselen also showed a strong influence on maintenance of the viability, morphology, and stem Cell function of the Cell sheets preserved under hypothermia by protecting them from reactive oxygen species-induced damage. Furthermore, ebselen drastically improved the preservation performance of human cornea tissues and their stem Cells. Therefore, ebselen shows good potential as a useful preservation agent in regenerative medicine as well as in cornea transplantation.
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transplantation of tissue Engineered epithelial Cell sheets after excimer laser photoablation reduces postoperative corneal haze
Investigative Ophthalmology & Visual Science, 2006Co-Authors: Yasutaka Hayashida, Joseph Yang, Kohji Nishida, K Watanabe, Akihiko Kikuchi, Naoyuki Maeda, Hiroaki Sugiyama, Yuichi Hori, Teruo OkanoAbstract:PURPOSE. To apply tissue-Engineered Cell sheet transplantation after excimer laser keratectomy as a novel approach for the reduction of postoperative comeal haze. METHODS. Limbal biopsy specimens were obtained, and epithelial Cells were cultured on temperature-responsive culture inserts without the use of feeder Cells. Laser keratectomy (7.0-mm ablation zone and 160-μm depth) was performed in the contralateral eye, and autologous epithelial Cell sheets were transplanted to the ablated comeal stroma. Transplant and control group eyes were assessed by slit lamp biomicroscopy, and comeal haze was scored in a masked fashion, according to the Fantes grading scale. For further examination histologic and immunohistochemical analyses were performed. RESULTS. Tissue-Engineered Cell sheets produced stable attachment to the laser-ablated sites, resulting in epithelialization, 5 minutes after transplantation. Conversely, control corneas required 3 to 5 days for complete re-epithelialization. At both 1 and 2 months after surgery, corneal haze was significantly inhibited in the transplant group. Histologic analyses showed that the number of keratocytes undergoing apoptosis was decreased in the transplant group at 3 days after surgery. Similarly, the expression of both collagen III and α-smooth muscle actin, which may enhance corneal haze, were diminished in the transplant group at 2 months. CONCLUSIONS. The transplantation of tissue-Engineered epithelial Cell sheets can successfully prevent the development of corneal haze after excimer laser keratectomy.
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corneal reconstruction with tissue Engineered Cell sheets composed of autologous oral mucosal epithelium
The New England Journal of Medicine, 2004Co-Authors: Kohji Nishida, Yasutaka Hayashida, K Watanabe, Kazuaki Yamamoto, Eijiro Adachi, Shigeru Nagai, Akihiko Kikuchi, Naoyuki Maeda, Hitoshi WatanabeAbstract:Background Ocular trauma or disease may lead to severe corneal opacification and, consequently, severe loss of vision as a result of complete loss of corneal epithelial stem Cells. Transplantation of autologous corneal stem-Cell sources is an alternative to allograft transplantation and does not require immunosuppression, but it is not possible in many cases in which bilateral disease produces total corneal stem-Cell deficiency in both eyes. We studied the use of autologous oral mucosal epithelial Cells as a source of Cells for the reconstruction of the corneal surface. Methods We harvested 3-by-3-mm specimens of oral mucosal tissue from four patients with bilateral total corneal stem-Cell deficiencies. Tissue-Engineered epithelial-Cell sheets were fabricated ex vivo by culturing harvested Cells for two weeks on temperature-responsive Cell-culture surfaces with 3T3 feeder Cells that had been treated with mitomycin C. After conjunctival fibrovascular tissue had been surgically removed from the ocular surfa...
Teruo Okano - One of the best experts on this subject based on the ideXlab platform.
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Improvement of the therapeutic capacity of insulin-producing Cells trans-differentiated from human liver Cells using Engineered Cell sheet
Stem Cell Research & Therapy, 2021Co-Authors: Yu Na Lee, Teruo Okano, Eun Hye Seo, Song Lee, Sarah Ferber, In Kyong Shim, Song Cheol KimAbstract:Background Although pancreatic islet transplantation therapy is ideal for diabetes patients, several hurdles have prevented it from becoming a standard treatment, including donor shortage and low engraftment efficacy. In this study, we prepared insulin-producing Cells trans-differentiated from adult human liver Cells as a new islet source. Also, Cell sheet formation could improve differentiation efficiency and graft survival. Methods Liver Cells were expanded in vitro and trans-differentiated to IPCs using adenovirus vectors carrying human genes for PDX1 , NEUROD1 , and MAFA . IPCs were seeded on temperature-responsive culture dishes to form Cell sheets. Differentiation efficiency was confirmed by ß Cell-specific gene expression, insulin production, and immunohistochemistry. IPC suspension was injected by portal vein (PV), and IPC sheet was transplanted on the liver surface of the diabetic nude mouse. The therapeutic effect of IPC sheet was evaluated by comparing blood glucose control, weight gain, histological evaluation, and hepatotoxicity with IPC injection group. Also, Cell biodistribution was assessed by in vivo/ex vivo fluorescence image tagging. Results Insulin gene expression and protein production were significantly increased on IPC sheets compared with those in IPCs cultured on conventional culture dishes. Transplanted IPC sheets displayed significantly higher engraftment efficiency and fewer transplanted Cells in other organs than injected IPCs, and also lower liver toxicity, improved blood glucose levels, and weight gain. Immunohistochemical analyses of liver tissue revealed positive staining for PDX1 and insulin at 1, 2, and 4 weeks after IPC transplantation. Conclusions In conclusion, Cell sheet formation enhanced the differentiation function and maturation of IPCs in vitro. Additionally, parameters for clinical application such as distribution, therapeutic efficacy, and toxicity were favorable. The Cell sheet technique may be used with IPCs derived from various Cell sources in clinical applications.
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middle ear mucosal regeneration by tissue Engineered Cell sheet transplantation
npj Regenerative Medicine, 2017Co-Authors: Kazuhisa Yamamoto, Masayuki Yamato, Ryo Takagi, Teruo Okano, Hiroaki Sugiyama, Tsunetaro Morino, Yuichiro Yaguchi, Hiromi KojimaAbstract:The recurrence of cholesteatoma after surgical treatment often occurs as a result of poor mucosal regeneration in the middle ear cavity and mastoid cavity and changes, such as granulation tissue formation, which impair gas exchange in the middle ear cavity. Conventional tympanoplasty often results in a lack of mucosal regeneration in the resected area of the mastoid cavity. In particular, mucosal regeneration in a poorly pneumatized mastoid cavity is extremely difficult. If the middle ear mucosa can be preserved or rapid postoperative regeneration of mucosa on the exposed bone surface can be achieved after middle ear surgery, the results of surgical treatment for otitis media, including cholesteatoma, can potentially be improved and the physiological function of the middle ear can be recovered. To overcome these limitations, we developed a novel treatment method combining tympanoplasty and autologous nasal mucosal epithelial Cell sheet transplantation for postoperative regeneration of the middle ear mucosa. In clinical research, we endoscopically removed an approximately 10 × 10 mm2 piece of nasal mucosal tissue. Tissue-Engineered autologous nasal mucosal epithelial Cell sheets were fabricated by culturing the harvested Cells in an aseptic environment in a good manufacturing practice-compliant Cell processing facility. The cultivated Cell sheets were transplanted, during tympanoplasty, onto the exposed bony surface of the attic of the tympanic and mastoid cavities where the mucosa had been lost. We performed this procedure on four patients with middle ear cholesteatoma and one patient with adhesive otitis media. All patients showed favorable postoperative course with no adverse events or complications and the patients’ hearing ability post-transplantation remained good. Transplanting a sheet of Cells from the nose to the middle ear could improve postoperative prognosis for middle ear inflammatory conditions. Kazuhisa Yamamoto of Jikei University School of Medicine and colleagues in Japan developed a novel surgical technique to prevent postoperative recurrence of chronic middle ear conditions. In adhesive otitis media, the eardrum gets sucked into and stuck in the middle ear space. Cholesteatoma involves abnormal skin growth in the middle ear. Recurrence often follows surgical treatment for these conditions due to delays in regeneration of the middle ear’s mucosal epithelium. The new technique involves transplanting cultured Cells from the lining of the nasal cavity to the middle ear during surgery. Five patients treated this way had favorable postoperative outcomes with no adverse effects or complications and satisfactory improvement in hearing levels.
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prevention of esophageal stricture after endoscopic submucosal dissection using tissue Engineered Cell sheets
Gastroenterology, 2012Co-Authors: Takeshi Ohki, Ryo Takagi, Teruo Okano, Daisuke Murakami, Makoto Kondo, Ryo Sasaki, Hideo Namiki, Masakazu YamamotoAbstract:Background & Aims The use of esophageal endoscopic submucosal dissection (ESD) to remove superficial esophageal neoplasms is gradually becoming more common in Japan. However, large-scale esophageal ESD often requires subsequent multiple balloon dilations to prevent postoperative esophageal stricture. We investigated the safety and efficacy of endoscopic transplantation of tissue-Engineered autologous oral mucosal epithelial Cell sheets in preventing formation of strictures after ESD. Methods We performed an open-label, single-arm, single-institute study. We collected specimens of oral mucosal tissue from 9 patients with superficial esophageal neoplasms. Epithelial Cell sheets were fabricated ex vivo by culturing isolated Cells for 16 days on temperature-responsive Cell culture surfaces. After a reduction in temperature, these sheets were endoscopically transplanted directly to the ulcer surfaces of patients who had just undergone ESD. All patients were monitored by endoscopy once a week until epithelialization was complete. Results Autologous Cell sheets were successfully transplanted to ulcer surfaces using an endoscope. Complete re-epithelialization occurred within a median time of 3.5 weeks. No patients experienced dysphagia, stricture, or other complications following the procedure, except for one patient who had a full circumferential ulceration that expanded to the esophagogastric junction. Conclusions Sutureless, endoscopic transplantation of carrier-free Cell sheets composed of autologous oral mucosal epithelial Cells safely and effectively promotes re-epithelialization of the esophagus after ESD. Patients in this study did not experience any serious complications. This procedure might be used to prevent stricture formation following ESD and improve patients' quality of life. Further study will be needed to show that stricture formation can be prevented.
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Functional closure of visceral pleural defects by autologous tissue Engineered Cell sheets
European Journal of Cardio-Thoracic Surgery, 2008Co-Authors: Masato Kanzaki, Masayuki Yamato, Joseph Yang, Hidekazu Sekine, Ryo Takagi, Tamami Isaka, Teruo Okano, Takamasa OnukiAbstract:Objective: The occurrence of intraoperative air leaks is an unavoidable complication during pulmonary surgeries. However, current surgical methodsaregenerallyineffective in closingthesevisceralpleuraldefects,resultingin a decreasedqualityoflife forpatients. Here,we examined novel tissue Engineered Cell sheets for the closure of pleural defects in a porcine model. Methods: Skin biopsies were harvested from juvenile swine and tissue sheets composed of dermal fibroblasts were created using ex vivo culture on temperature-responsive dishes. After creating a visceral pleuralinjurymodel, thetissue Engineeredautologousdermalfibroblastsheetswere transplanteddirectlyto the defectswithoutthe use of sutures or additional adhesive agents, such as fibrin glue. Results: The tissue Engineered autologous dermal fibroblast sheets attached directly to the lung surface providing an immediate seal against up to 25 cm H2O of airway pressure. Four weeks after transplantation, the dermal fibroblast sheets remained present on the pleural surface, providing permanent closure. The dermal fibroblast sheets were also responsive to changes in lung volume due to mechanical ventilation. No recurrences of air leaks were observed throughout the follow-up period. Conclusions: This study presents the development of an effective sealant for visceral pleural defects using autologous Cells that have the flexibility to respond to expansion and contraction during respiration. # 2008 European Association for Cardio-Thoracic Surgery. Published by Elsevier B.V. All rights reserved.
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Dynamic sealing of lung air leaks by the transplantation of tissue Engineered Cell sheets
Biomaterials, 2007Co-Authors: Masato Kanzaki, Joseph Yang, Hidekazu Sekine, Chinatsu Kohno, Ryo Takagi, Hideyuki Hatakeyama, Tamami Isaka, Teruo Okano, Takamasa OnukiAbstract:Current methods including the use of various biological and synthetic sealants are ineffective in the closure of intraoperative air leaks that often occur during cardiothoracic surgeries, resulting in a decreased quality of life for patients. We present the development of a novel lung air leak sealant using tissue Engineered Cell sheets. In contrast to previous materials such as fibrin glue, these bioEngineered Cell sheets immediately and permanently seal air leaks in a dynamic fashion that allows for the extensive tissue contraction and expansion involved in respiration, without any postoperative recurrences. Additionally, we demonstrate that mesothelial Cells migrate to cover the transplanted Cells sheets, thereby confirming exCellent biocompatibility and integration with the host tissues. Finally, we present the use of skin fibroblasts as an effective and readily available autologous Cell source that can be easily applied. This study shows for the first time, the development of an immediate and permanent lung air leak sealant, suitable for future clinical applications.
Charlie Y.m. Hsu - One of the best experts on this subject based on the ideXlab platform.
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An Integrated Approach toward the Biomanufacturing of Engineered Cell Therapy Products in a Stirred-Suspension Bioreactor.
Molecular therapy. Methods & clinical development, 2018Co-Authors: Charlie Y.m. Hsu, Tylor Walsh, Breanna S. Borys, Michael S. Kallos, Derrick E. RancourtAbstract:Recent advances in stem Cell biology have accelerated the pre-clinical development of Cell-based therapies for degenerative and chronic diseases. The success of this growing area hinges upon the concomitant development of scalable manufacturing platforms that can produce clinically relevant quantities of Cells for thousands of patients. Current biomanufacturing practices for Cell therapy products are built on a model previously optimized for biologics, wherein stable Cell lines are established first, followed by large-scale production in the bioreactor. This "two-step" approach can be costly, labor-intensive, and time-consuming, particularly for Cell therapy products that must be individually sourced from patients or compatible donors. In this report, we describe a "one-step" integrated approach toward the biomanufacturing of Engineered Cell therapy products by direct transfection of primary human fibroblast in a continuous stirred-suspension bioreactor. We optimized the transfection efficiency by testing rate-limiting factors, including Cell seeding density, agitation rate, oxygen saturation, microcarrier type, and serum concentration. By combining the genetic modification step with the large-scale expansion step, this not only removes the need for manual handing of Cells in planar culture dishes, but also enables the biomanufacturing process to be streamlined and automated in one fully enclosed bioreactor.
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An Integrated Approach toward the Biomanufacturing of Engineered Cell Therapy Products in a Stirred-Suspension Bioreactor
Elsevier, 2018Co-Authors: Charlie Y.m. Hsu, Tylor Walsh, Breanna S. Borys, Michael S. Kallos, Derrick E. RancourtAbstract:Recent advances in stem Cell biology have accelerated the pre-clinical development of Cell-based therapies for degenerative and chronic diseases. The success of this growing area hinges upon the concomitant development of scalable manufacturing platforms that can produce clinically relevant quantities of Cells for thousands of patients. Current biomanufacturing practices for Cell therapy products are built on a model previously optimized for biologics, wherein stable Cell lines are established first, followed by large-scale production in the bioreactor. This “two-step” approach can be costly, labor-intensive, and time-consuming, particularly for Cell therapy products that must be individually sourced from patients or compatible donors. In this report, we describe a “one-step” integrated approach toward the biomanufacturing of Engineered Cell therapy products by direct transfection of primary human fibroblast in a continuous stirred-suspension bioreactor. We optimized the transfection efficiency by testing rate-limiting factors, including Cell seeding density, agitation rate, oxygen saturation, microcarrier type, and serum concentration. By combining the genetic modification step with the large-scale expansion step, this not only removes the need for manual handing of Cells in planar culture dishes, but also enables the biomanufacturing process to be streamlined and automated in one fully enclosed bioreactor. Keywords: bioprocess, gene delivery, transfection, biotechnology, cationic polymer, Cell therapy, biomanufacturing, genetic engineering, plasmid DNA, bioreacto
Tylor Walsh - One of the best experts on this subject based on the ideXlab platform.
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An Integrated Approach toward the Biomanufacturing of Engineered Cell Therapy Products in a Stirred-Suspension Bioreactor.
Molecular therapy. Methods & clinical development, 2018Co-Authors: Charlie Y.m. Hsu, Tylor Walsh, Breanna S. Borys, Michael S. Kallos, Derrick E. RancourtAbstract:Recent advances in stem Cell biology have accelerated the pre-clinical development of Cell-based therapies for degenerative and chronic diseases. The success of this growing area hinges upon the concomitant development of scalable manufacturing platforms that can produce clinically relevant quantities of Cells for thousands of patients. Current biomanufacturing practices for Cell therapy products are built on a model previously optimized for biologics, wherein stable Cell lines are established first, followed by large-scale production in the bioreactor. This "two-step" approach can be costly, labor-intensive, and time-consuming, particularly for Cell therapy products that must be individually sourced from patients or compatible donors. In this report, we describe a "one-step" integrated approach toward the biomanufacturing of Engineered Cell therapy products by direct transfection of primary human fibroblast in a continuous stirred-suspension bioreactor. We optimized the transfection efficiency by testing rate-limiting factors, including Cell seeding density, agitation rate, oxygen saturation, microcarrier type, and serum concentration. By combining the genetic modification step with the large-scale expansion step, this not only removes the need for manual handing of Cells in planar culture dishes, but also enables the biomanufacturing process to be streamlined and automated in one fully enclosed bioreactor.
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An Integrated Approach toward the Biomanufacturing of Engineered Cell Therapy Products in a Stirred-Suspension Bioreactor
Elsevier, 2018Co-Authors: Charlie Y.m. Hsu, Tylor Walsh, Breanna S. Borys, Michael S. Kallos, Derrick E. RancourtAbstract:Recent advances in stem Cell biology have accelerated the pre-clinical development of Cell-based therapies for degenerative and chronic diseases. The success of this growing area hinges upon the concomitant development of scalable manufacturing platforms that can produce clinically relevant quantities of Cells for thousands of patients. Current biomanufacturing practices for Cell therapy products are built on a model previously optimized for biologics, wherein stable Cell lines are established first, followed by large-scale production in the bioreactor. This “two-step” approach can be costly, labor-intensive, and time-consuming, particularly for Cell therapy products that must be individually sourced from patients or compatible donors. In this report, we describe a “one-step” integrated approach toward the biomanufacturing of Engineered Cell therapy products by direct transfection of primary human fibroblast in a continuous stirred-suspension bioreactor. We optimized the transfection efficiency by testing rate-limiting factors, including Cell seeding density, agitation rate, oxygen saturation, microcarrier type, and serum concentration. By combining the genetic modification step with the large-scale expansion step, this not only removes the need for manual handing of Cells in planar culture dishes, but also enables the biomanufacturing process to be streamlined and automated in one fully enclosed bioreactor. Keywords: bioprocess, gene delivery, transfection, biotechnology, cationic polymer, Cell therapy, biomanufacturing, genetic engineering, plasmid DNA, bioreacto