共查询到20条相似文献,搜索用时 15 毫秒
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Joanna Filipowska Gwendolen C. Reilly Anna M. Osyczka 《Biotechnology and bioengineering》2016,113(8):1814-1824
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Scaglione S Braccini A Wendt D Jaquiery C Beltrame F Quarto R Martin I 《Biotechnology and bioengineering》2006,93(1):181-187
In this work, we investigated whether osteoinductive constructs can be generated by isolation and expansion of sheep bone marrow stromal cells (BMSC) directly within three-dimensional (3D) ceramic scaffolds, bypassing the typical phase of monolayer (2D) expansion prior to scaffold loading. Nucleated cells from sheep bone marrow aspirate were seeded into 3D ceramic scaffolds either by static loading or under perfusion flow and maintained in culture for up to 14 days. The resulting constructs were exposed to enzymatic treatment to assess the number and lineage of extracted cells, or implanted subcutaneously in nude mice to test their capacity to induce bone formation. As a control, BMSC expanded in monolayer for 14 days were also seeded into the scaffolds and implanted. BMSC could be isolated and expanded directly in the 3D ceramic scaffolds, although they proliferated slower than in 2D. Upon ectopic implantation, the resulting constructs formed a higher amount of bone tissue than constructs loaded with the same number of 2D-expanded cells. Constructs cultivated for 14 days generated significantly more bone tissue than those cultured for 3 days. No differences in bone formation were found between samples seeded by static loading or under perfusion. In conclusion, the culture of bone marrow nucleated cells directly on 3D ceramic scaffolds represents a promising approach to expand BMSC and streamline the engineering of osteoinductive grafts. 相似文献
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Perfusion bioreactor systems play a crucial role in mitigating nutrient limitation as well as providing biomechanical stimuli and redistributing regulatory macromolecules that influence human mesenchymal stem cells (hMSC) fate in three‐dimensional (3D) scaffolds. As fibroblast growth factor‐2 (FGF‐2) is known to regulate hMSC phenotype, understanding the role of autocrine FGF‐2 signaling in the 3D construct under the different perfusion flow provides important insight into an optimal bioreactor design. To investigate FGF‐2 signaling inhibition in hMSC cultured in the porous poly(ethylene terephthalate) (PET) scaffolds perfused under two flow configurations, PD173074, an FGFR1 inhibitor, was added in growth media after 7 day of pre‐culture and its impact on hMSC proliferation and clonogenicity during the subsequent 7 days of cultivation was analyzed. Compared with control constructs in growth media, the addition of PD173074 resulted in significant reduction in hMSC proliferation and colony formation in both constructs with a more dramatic reduction in the parallel flow constructs. The results demonstrate that autocrine FGF‐2 plays a significant role in 3D scaffold and suggest modulation of the perfusion flow in the bioreactor as a strategy to influence autocrine actions and cell fate in the 3D scaffold. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012 相似文献
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Celeste Manfredonia Manuele G. Muraro Christian Hirt Valentina Mele Valeria Governa Adam Papadimitropoulos Silvio Dster Savas D. Soysal Raoul A. Droeser Robert Mechera Daniel Oertli Raffaele Rosso Martin Bolli Andreas Zettl Luigi M. Terracciano Giulio C. Spagnoli Ivan Martin Giandomenica Iezzi 《Advanced Biosystems》2019,3(4)
Colorectal cancer (CRC) is a leading cause of cancer‐related death. Conventional chemotherapeutic regimens have limited success rates, and a major challenge for the development of novel therapies is the lack of adequate in vitro models. Nonmalignant mesenchymal and immune cells of the tumor microenvironment (TME) are known to critically affect CRC progression and drug responsiveness. However, tumor drug sensitivity is still evaluated on systems, such as cell monolayers, spheroids, or tumor xenografts, which typically neglect the original TME. Here, it is investigated whether a bioreactor‐based 3D culture system can preserve the main TME cellular components in primary CRC samples. Freshly excised CRC fragments are inserted between two collagen scaffolds in a “sandwich‐like” format and cultured under static or perfused conditions up to 3 d. Perfused cultures maintain tumor tissue architecture and densities of proliferating tumor cells to significantly higher extents than static cultures. Stromal and immune cells are also preserved and fully viable, as indicated by their responsiveness to microenvironmental stimuli. Importantly, perfusion‐based cultures prove suitable for testing the sensitivity of primary tumor cells to chemotherapies currently in use for CRC. Perfusion‐based culture of primary CRC specimens recapitulates TME key features and may allow assessment of tumor drug response in a patient‐specific context. 相似文献
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Shear stress is an important biomechanical parameter in regulating human mesenchymal stem cell (hMSC) construct development. In this study, the biomechanical characteristics of hMSCs within highly porous 3-D poly (ethylene terephthalate) (PET) matrices in a perfusion bioreactor system were analyzed for two flow rates of 0.1 and 1.5 mL/min, respectively over a 20-day culture period. A 1.4 times higher proliferation rate, higher CFU-F formation, and more fibronectin and HSP-47 secretion at day 20 were observed at the flow rate of 0.1 mL/min compared to those at the flow rate of 1.5 mL/min. The higher flow rate of 1.5 mL/min upregulated osteogenic differentiation potential at day 20 as measured by the expression of alkaline phosphatase activity and calcium deposition in the matrix after 14 days osteogenic induction, consistent with those reported in literatures. Mathematical modeling indicated that shear stress existed in the range of 1 x 10(-5) to 1 x 10(-4) Pa in the constructs up to a depth of 70 microm due to flow penetration in the porous constructs. Analysis of oxygen transport in the constructs for the two flow rates yielded oxygen levels significantly higher than those at which cell growth and metabolism are affected (Jiang et al., 1996). This indicates that differences in convective transport have no significant influence on cell growth and metabolism for the range of flow rates studied. These results demonstrate that shear stress is an important microenvironment parameter that regulates hMSC construct development at a range significantly lower than those reported previously in the perfusion system. 相似文献
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Alyaa I. Aldebs Fatema T. Zohora Nasim Nosoudi Surinder P. Singh Jaime E. Ramirez-Vick 《Bioelectromagnetics》2020,41(3):175-187
Alternative bone regeneration strategies that do not rely on harvested tissue or exogenous growth factors are needed. One of the major challenges in tissue reconstruction is recreating the bone tissue microenvironment using the appropriate combination of cells, scaffold, and stimulation to direct differentiation. This study presents a bone regeneration formulation that involves the use of human adipose-derived mesenchymal stem cells (hASCs) and a three-dimensional (3D) hydrogel scaffold based on self-assembled RADA16 peptides containing superparamagnetic iron oxide nanoparticles (NPs). Although superparamagnetic NPs could be used as stimulus to manipulate the cell proliferation and differentiation, in this paper their use is explored for assisting osteogenic differentiation of hASCs in conjunction with direct stimulation by extremely low-frequency pulsed electromagnetic fields (pEMFs). Cellular morphology, proliferation, and viability, as well as alkaline phosphatase activity, calcium deposition, and osteogenic capacity were monitored for cells cultured up to 21 days in the 3D construct. The results show that the pEMFs and NPs do not have any negative effect on cell viability, but instead distinctly induced early differentiation of hASCs to an osteoblastic phenotype, when compared with cells without biophysical stimulation. This effect is attributed to synergy between the pEMFs and NPs, which may have stimulated mechanotransduction pathways, which, in turn activated biochemical signals between cells to differentiate or proliferate. This approach may offer a safe and effective option for the treatment of non-union bone fractures. Bioelectromagnetics. © 2020 The Authors. Bioelectromagnetics published by Wiley Periodicals, Inc. 相似文献
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间充质干细胞(mesenchyrmalstemcells,MSCs)是当前在多种组织再生和细胞治疗研究中被最广泛采用的一类干细胞。但如何诱导MSCs的体外高效扩增并维持其干性特征(stemness),从而为临床应用提供充足、优质的细胞源,是当前基础研究和临床治疗中遇到的瓶颈问题。日益增多的研究表明,机体内干细胞的自我更新与分化受其所处体内微环境的紧密调控。因此,精确模拟干细胞在体内生长的微环境已成为提高干细胞体外扩增效率的重要策略。该文就近期研究中如何模拟干细胞生长微环境诱导MSCs体外扩增并维持干细胞特性的研究做一综述,为今后MSCs的高效扩增和推进临床运用与转化提供思路。 相似文献
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Human mesenchymal stem cells (hMSCs) have great potential for therapeutic applications. A bioreactor system that supports long-term hMSCs growth and three-dimensional (3-D) tissue formation is an important technology for hMSC tissue engineering. A 3-D perfusion bioreactor system was designed using non-woven poly (ethylene terepthalate) (PET) fibrous matrices as scaffolds. The main features of the perfusion bioreactor system are its modular design and integrated seeding operation. Modular design of the bioreactor system allows the growth of multiple engineered tissue constructs and provides flexibility in harvesting the constructs at different time points. In this study, four chambers with three matrices in each were utilized for hMSC construct development. The dynamic depth filtration seeding operation is incorporated in the system by perfusing cell suspensions perpendicularly through the PET matrices, achieving a maximum seeding efficiency of 68%, and the operation effectively reduced the complexity of operation and the risk of contamination. Statistical analyses suggest that the cells are uniformly distributed in the matrices. After seeding, long-term construct cultivation was conducted by perfusing the media around the constructs from both sides of the matrices. Compared to the static cultures, a significantly higher cell density of 4.22 x 10(7) cell/mL was reached over a 40-day culture period. Cellular constructs at different positions in the flow chamber have statistically identical cell densities over the culture period. After expansion, the cells in the construct maintained the potential to differentiate into osteoblastic and adipogenic lineages at high cell density. The perfusion bioreactor system is amenable to multiple tissue engineered construct production, uniform tissue development, and yet is simple to operate and can be scaled up for potential clinical use. The results also demonstrate that the multi-lineage differentiation potential of hMSCs are preserved even after extensive expansion, thus indicating the potential of hMSCs for functional tissue construct development. The system has important applications in stem cell tissue engineering. 相似文献
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Zahra Jamalpoor Mansoureh Soleimani Nafise Taromi Alireza Asgari 《Journal of cellular physiology》2019,234(12):23123-23134
Expansion of seeded human Wharton's jelly mesenchymal stem cells (hWJ-MSCs) on 2D culture plates and 3D nano-hydroxyapatite/chitosan/gelatin scaffolds, from morphology and osteoactivity points of view, were investigated. Cell attachment and spreading, temporal expression profiles of selected osteogenic gene and protein markers, intracellular alkaline phosphatase enzyme activity (ALP activity), and matrix mineralization were assayed over the course of the experiments. Morphological results demonstrated hWJ-MSCs had greater affinity to adhere onto the 3D scaffold surface, as the number and thickness of the filopodia were higher in the 3D compared with 2D culture system. Functionally, the intracellular ALP activity and extracellular mineralization in 3D scaffolds were significantly greater, in parallel with elevation of osteogenic markers at the mRNA and protein levels at all-time point. It is concluded that 3D scaffolds, more so than 2D culture plate, promote morphology and osteogenic behavior of WJ-MSCs in vitro, a promising system for MSCs expansion without compromising their stemness before clinical transplantation. 相似文献
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Reliably producing functional in vitro organ models, such as organ-on-chip systems, has the potential to considerably advance biology research, drug development time, and resource efficiency. However, despite the ongoing major progress in the field, three-dimensional bone tissue models remain elusive. In this review, we specifically investigate the control of perfusion flow effects as the missing link between isolated culture systems and scientifically exploitable bone models and propose a roadmap toward this goal. 相似文献
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Scheufler O Schaefer DJ Jaquiery C Braccini A Wendt DJ Gasser JA Galli R Pierer G Heberer M Martin I 《Journal of cellular and molecular medicine》2008,12(4):1238-1249
Biological substitutes for autologous bone flaps could be generated by combining flap pre-fabrication and bone tissue engineering concepts. Here, we investigated the pattern of neotissue formation within large pre-fabricated engineered bone flaps in rabbits. Bone marrow stromal cells from 12 New Zealand White rabbits were expanded and uniformly seeded in porous hydroxyapatite scaffolds (tapered cylinders, 10-20 mm diameter, 30 mm height) using a perfusion bioreactor. Autologous cell-scaffold constructs were wrapped in a panniculus carnosus flap, covered by a semipermeable membrane and ectopically implanted. Histological analysis, substantiated by magnetic resonance imaging (MRI) and micro-computerized tomography scans, indicated three distinct zones: an outer one, including bone tissue; a middle zone, formed by fibrous connective tissue; and a central zone, essentially necrotic. The depths of connective tissue and of bone ingrowth were consistent at different construct diameters and significantly increased from respectively 3.1 +/- 0.7 mm and 1.0 +/- 0.4 mm at 8 weeks to 3.7+/- 0.6 mm and 1.4 +/- 0.6 mm at 12 weeks. Bone formation was found at a maximum depth of 1.8 mm after 12 weeks. Our findings indicate the feasibility of ectopic pre-fabrication of large cell-based engineered bone flaps and prompt for the implementation of strategies to improve construct vascularization, in order to possibly accelerate bone formation towards the core of the grafts. 相似文献
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Samaneh Shojaei Seyed Mahmoud Hashemi Hossein Ghanbarian Mohammad Salehi Samira Mohammadi-Yeganeh 《Journal of cellular physiology》2019,234(4):3394-3409
Mesenchymal stem cells (MSCs) are multipotent cells with the potential to differentiate into different cell types. Owing to their immunosuppressive and anti-inflammatory properties, they are widely used in regenerative medicine, but they have a dual effect on cancer progression and exert both growth-stimulatory or -inhibitory effects on different cancer types. It has been proposed that these controversial effects of MSC in tumor microenvironment (TME) are mediated by their polarization to proinflammatory or anti-inflammatory phenotype. In addition, they can polarize the immune system cells that in turn influence tumor progression. One of the mechanisms involved in the TME communications is extracellular vesicles (EVs). MSCs, as one of cell populations in TME, produce a large amount of EVs that can influence tumor development. Similar to MSC, MSC-EVs can exert both anti- or protumorigenic effects. In the current study, we will investigate the current knowledge related to MSC role in cancer progression with a focus on the MSC-EV content in limiting tumor growth, angiogenesis, and metastasis. We suppose MSC-EVs can be used as safe vehicles for delivering antitumor agents to TME. 相似文献
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Polypeptide changes associated with loss of proliferative potential during the terminal event in differentiation 总被引:2,自引:0,他引:2
The differentiation of murine mesenchymal stem cells occurs in nonterminal and terminal phases. In previous reports we established the characteristics of nonterminally differentiated cells and showed that transition from the nonterminal to the terminal state of differentiation can be induced by human plasma. We also showed that this transition is blocked by protein synthesis inhibitors and other pharmacological agents. In this paper, we have employed two-dimensional gel electrophoresis to evaluate changes in specific polypeptides that are induced when cells lose proliferative capacity associated with the terminal event in differentiation. Using silver staining procedures for analysis of electrophoretograms, we detected only seven major polypeptide differences between nonterminally differentiated and terminally differentiated cells. Six polypeptides were expressed only in preparations of terminally differentiated cells; these included two polypeptides identified in cytosolic fractions and four polypeptides identified in nuclear fractions. One polypeptide was also found to be selectively expressed only in nuclear fractions of nonterminally differentiated cells. Based on these observations we conclude that the loss of proliferative potential that occurs during the terminal event in mesenchymal stem cell differentiation is associated with changes in the composition of a limited number of specific polypeptides. We suggest that one or more of these polypeptides may be important in the regulation of cellular proliferation. 相似文献
19.
The characteristics and multilineage differentiation potential of bone marrow mesenchymal stem cells (BM MSC) remain controversial. This study aimed to characterize human BM MSC isolated by plastic adherent or antibody selection and their neuronal differentiation potential using growth factors or chemical inducing agents. MSC were found to express low levels of neuronal markers: neurofilament-M, beta tubulin III, and neuron specific enolase. Under a serum- and feeder cell-free condition, basic fibroblast growth factor, epidermal growth factor, and platelet-derived growth factor induced neuronal morphology in MSC. In addition to the above markers, these cells expressed neurotransmitters or associated proteins: gamma-aminobutyric acid, tyrosine hydroxylase and serotonin. These changes were maintained for up to 3 months in all bone marrow specimens (N = 6). In contrast, butylated hydroxyanisole and dimethylsulfoxide were unable to induce sustained neuronal differentiation. Our results show that MSC isolated by two different procedures produced identical lineage differentiation with defined growth factors in a serum- and feeder cell-free condition. 相似文献
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Teng Ma Warren L. Grayson Mirjam Fröhlich Gordana Vunjak‐Novakovic 《Biotechnology progress》2009,25(1):32-42
Stem cells have the ability for prolonged self‐renewal and differentiation into mature cells of various lineages, which makes them important cell sources for tissue engineering applications. Their remarkable ability to replenish and differentiate in vivo is regulated by both intrinsic and extrinsic cellular mechanisms. The anatomical location where the stem cells reside, known as the “stem cell niche or microenvironment,” provides signals conducive to the maintenance of definitive stem cell properties. Physiological condition including oxygen tension is an important component of the stem cell microenvironment and has been shown to play a role in regulating both embryonic and adult stem cells. This review focuses on oxygen as a signaling molecule and the way it regulates the stem cells' development into mesenchymal tissues in vitro. The physiological relevance of low oxygen tension as an environmental parameter that uniquely benefits stem cells' expansion and maintenance is described along with recent findings on the regulatory effects of oxygen on embryonic stem cells and adult mesenchymal stem cells. The relevance to tissue engineering is discussed in the context of the need to specifically regulate the oxygen content in the cellular microenvironment in order to optimize in vitro tissue development. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009 相似文献