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1.
Cytotherapy ; 20(6): 779-784, 2018 06.
Article in English | MEDLINE | ID: mdl-29784433

ABSTRACT

The development of standards for the field of regenerative medicine has been noted as a high priority by several road-mapping activities. Additionally, the U.S. Congress recognizes the importance of standards in the 21st Century Cure Act. Standards will help to accelerate and streamline cell and gene therapy product development, ensure the quality and consistency of processes and products, and facilitate their regulatory approval. Although there is general agreement for the need of additional standards for regenerative medicine products, a shared understanding of standards is required for real progress toward the development of standards to advance regenerative medicine. Here, we describe the roles of standards in regenerative medicine as well as the process for standards development and the interactions of different entities in the standards development process. Highlighted are recent coordinated efforts between the U.S. Food and Drug Administration and the National Institute of Standards and Technology to facilitate standards development and foster science that underpins standards development.


Subject(s)
Biological Products/standards , Cooperative Behavior , Inventions/standards , Regenerative Medicine/standards , Therapies, Investigational/standards , Translational Research, Biomedical/standards , United States Food and Drug Administration , Biological Products/therapeutic use , Drug Approval , Genetic Therapy/methods , Genetic Therapy/standards , Genetic Therapy/trends , Humans , Intersectoral Collaboration , Inventions/trends , Reference Standards , Regenerative Medicine/methods , Regenerative Medicine/organization & administration , Therapies, Investigational/methods , Translational Research, Biomedical/methods , Translational Research, Biomedical/organization & administration , United States
2.
Acta Biomater ; 32: 161-169, 2016 Mar 01.
Article in English | MEDLINE | ID: mdl-26773464

ABSTRACT

The interactions between cells and an underlying biomaterial are important for the promotion of cell adhesion, proliferation, and function. Mesenchymal stem cells (MSCs) have great clinical potential as they are an adult stem cell population capable of multilineage differentiation. The relationship between MSC behavior and several material properties including substrate stiffness and pore size are well investigated, but there has been little research on the influence of porous architecture in a three-dimensional scaffold with a well-controlled architecture. Here, we investigate the impact of two different three-dimensionally printed, pore geometries on the enrichment and differentiation of MSCs. 3D printed scaffolds with ordered cubic pore geometry were supportive of MSC enrichment from unprocessed bone marrow, resulting in cell surface marker expression that was comparable to typical adhesion to tissue culture polystyrene, the gold standard for MSC culture. Results also show that scaffolds fabricated with ordered cubic pores significantly increase the gene expression of MSCs undergoing adipogenesis and chondrogenesis, when compared to scaffolds with ordered cylindrical pores. However, at the protein expression level, these differences were modest. For MSCs undergoing osteogenesis, gene expression results suggest that cylindrical pores may initially increase early osteogenic marker expression, while protein level expression at later timepoints is increased for scaffolds with ordered cubic pores. Taken together, these results suggest that 3D printed scaffolds with ordered cubic pores could be a suitable culture system for single-step MSC enrichment and differentiation. STATEMENT OF SIGNIFICANCE: Mesenchymal stem cells (MSCs) have great therapeutic potential, as they are capable of multilineage differentiation. MSC behavior, including lineage commitment, may be influenced by biomaterial properties including substrate stiffness and pore size. With three-dimensional (3D) printing, we can investigate these relationships in 3D culture systems. Here, we fabricated scaffolds with two different well-controlled pore geometries, and investigated the impact on MSC enrichment and differentiation. Results show that scaffolds with ordered cubic pore geometry were supportive of both MSC enrichment from unprocessed bone marrow as well as MSC differentiation, resulting in increased gene expression during adipogenesis and chondrogenesis. These results suggest that 3D printed scaffolds with ordered cubic pores could be a suitable culture system for single-step MSC enrichment and differentiation.


Subject(s)
Cell Differentiation , Mesenchymal Stem Cells/cytology , Printing, Three-Dimensional , Biomarkers/metabolism , Cell Differentiation/genetics , Cell Proliferation , Cell Survival , Flow Cytometry , Gene Expression Regulation , Humans , Porosity , Tissue Scaffolds
3.
Tissue Eng Part C Methods ; 22(4): 348-59, 2016 Apr.
Article in English | MEDLINE | ID: mdl-26797048

ABSTRACT

Despite great promise surrounding mesenchymal stem cells (MSCs), their implementation for tissue engineering strategies remains in the development phases. Many of the concerns regarding the clinical use of MSCs originate from population heterogeneity, during both isolation and differentiation. In this study, we utilize our previously developed centrifugation cell adhesion protocol for the separation of MSCs. Our findings reveal that MSCs can be isolated from whole bone marrow using a 200 g (700 pN) centrifugal force after 24 h of culture on polystyrene with cell surface marker expression equivalent to positive controls. During differentiation, a centrifugation protocol with identical force parameters could be applied 14 days into chondrogenic differentiation to isolate differentiated chondrocytes, which exhibited increased expression of chondrogenic markers compared to controls. In summary, the use of our developed centrifugation cell adhesion protocol has proven to be an effective means to separate MSC populations, decreasing the heterogeneity of subsequent cell therapy products.


Subject(s)
Antigens, Differentiation/biosynthesis , Bone Marrow Cells , Cell Differentiation , Cell Separation/methods , Chondrogenesis , Mesenchymal Stem Cells , Bone Marrow Cells/cytology , Bone Marrow Cells/metabolism , Centrifugation , Humans , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/metabolism
4.
Mol Pharm ; 11(7): 2172-81, 2014 Jul 07.
Article in English | MEDLINE | ID: mdl-24620713

ABSTRACT

The importance of providing a physiologically relevant environment for cell culture is well recognized. The combination of proper environmental cues which are provided in vivo by the bloodstream and extracellular matrix must be reproduced to properly examine cell response in vitro, and cannot be recapitulated using traditional culture on polystyrene. Here, we have developed a device, the dynamic stem cell culture platform (DSCCP), consisting of a biomimetic scaffold cultured within the dynamic environment of a perfusion bioreactor. By varying scaffold parameters including stiffness and protein inclusion at the material surface, we found that human mesenchymal stem cells (hMSCs) were able to adhere to modified substrates, while still maintaining multipotency. Culture in a perfusion bioreactor showed cell survival and proliferation, particularly on modified substrates. The DSCCP represents a complete platform for cell adhesion and subsequent evaluation, including the response of a cell population to drug treatment.


Subject(s)
Cell Adhesion/physiology , Cell Culture Techniques/methods , Mesenchymal Stem Cells/physiology , Biomimetics/methods , Bioreactors , Cell Differentiation/physiology , Cell Proliferation/physiology , Cell Survival/physiology , Extracellular Matrix/physiology , Humans , Perfusion/methods , Tissue Engineering/methods , Tissue Scaffolds
5.
Tissue Eng Part C Methods ; 18(7): 537-44, 2012 Jul.
Article in English | MEDLINE | ID: mdl-22235797

ABSTRACT

A major obstacle in chondrocyte-based therapy for cartilage repair is the limited availability of cells that maintain their original phenotype. Propagation of chondrocytes as monolayer cultures on polystyrene surfaces is used extensively for amplifying cell numbers. However, chondrocytes undergo a phenotypic shift when propagated in this manner and display characteristics of more adherent fibroblastic cells. Little information is available about the effect of this phenotypic shift on cellular adhesion properties. We evaluated changes in adhesion property as bovine chondrocytes were serially propagated up to five passages in monolayer culture using a centrifugation cell adhesion assay, which was based on counting of cells before and after being exposed to centrifugal dislodgement forces of 120 and 350 g. Chondrocytes proliferated well in a monolayer culture with doubling times of 2-3 days, but they appeared more fibroblastic and exhibited elongated cell morphology with continued passage. The centrifugation cell adhesion assay showed that chondrocytes became more adhesive with passage as the percentage of adherent cells after centrifugation increased and was not statistically different from the adhesion of the fibroblast cell line, L929, starting at passage 3. This increased adhesiveness correlated with a shift to a fibroblastic morphology and increased collagen I mRNA expression starting at passage 2. Our findings indicate that the centrifugation cell adhesion assay may serve as a reproducible tool to track alterations in chondrocyte phenotype during their extended propagation in culture.


Subject(s)
Cell Adhesion/physiology , Cell Culture Techniques , Centrifugation , Chondrocytes/cytology , Chondrocytes/metabolism , Polystyrenes/metabolism , Animals , Blotting, Western , Cattle , Cell Differentiation , Cell Proliferation , Cells, Cultured , Collagen Type II/genetics , Collagen Type II/metabolism , Fibroblasts/cytology , Fibroblasts/metabolism , Immunoenzyme Techniques , Mice , Phenotype , RNA, Messenger/genetics , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction , Serial Passage
7.
Article in English | MEDLINE | ID: mdl-16163399

ABSTRACT

BACKGROUND: This open-label pilot study investigated whether the selective serotonin reuptake inhibitor (SSRI) citalopram improves symptoms of irritable bowel syndrome (IBS), a functional gastrointestinal disorder with frequent psychiatric comorbidity. METHOD: Fifteen patients meeting Rome I criteria for IBS were administered open-label citalopram (20-40 mg/day) for 12 weeks. The study was conducted from October 2000 to August 2001. RESULTS: Twelve (80%) of the 15 subjects reported a > or = 50% decrease in the presence of abdominal pain, 10 (67%) reported a > or = 50% reduction in the severity of the symptom, and 12 (80%) reported a > or = 50% reduction in the frequency of the symptom. Approximately one half of the patients met criteria for remission (> or = 70% improvement) of abdominal pain. CONCLUSION: Results of this pilot study suggest that large controlled trials are needed to further evaluate the efficacy of SSRIs such as citalopram for the treatment of IBS.

8.
Article in English | MEDLINE | ID: mdl-15014729

ABSTRACT

BACKGROUND: Irritable bowel syndrome (IBS) is a common disorder and is the largest diagnostic cohort seen by gastroenterologists. There is a bidirectional comorbidity of IBS and psychiatric illness. Ours is the first study to examine the effect of any selective serotonin reuptake inhibitor in subjects with IBS. METHOD: Twenty subjects with Rome I criteria-diagnosed IBS were treated with 20 to 40 mg of paroxetine for 12 weeks. We utilized a computer-administered patient daily questionnaire taken by patients over the telephone using an interactive voice response system. RESULTS: Sixty-five percent of patients (13/20) reported a reduction in abdominal pain, and 55% (11/20) reported a reduction in pain frequency (total or mean number of days per week in which the patient had the symptom decreased by >/= 50%). Constipation and diarrhea were reduced in 69% and 57% of patients (9/13 and 8/14), respectively. Similarly, a clinically significant reduction in the symptoms of feeling of incomplete emptying (53% [9/17]) and bloating/abdominal distension (55% [11/20]) was apparent at study conclusion compared with baseline. On the Clinical Global Impressions scale at week 12, 47% (8/17) of the patients were much or very much improved. CONCLUSION: In our pilot open-label study, paroxetine was very effective in alleviating the abdominal pain and associated symptoms of IBS. These results warrant further examination in a placebo-controlled study.

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