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RISS 인기검색어
- 검색키워드
- 저자 : Behrad Vahidi (검색결과 3 건)
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Quantitative Analysis of CNS Axon Regeneration Using a Microfluidic Neuron Culture Device
박정원,Behrad Vahidi,김형준,이석우,전누리 한국바이오칩학회 2008 BioChip Journal Vol.2 No.1
CNS axonal regeneration has been widely studied in order to develop strategies for overcoming Myelin mediated inhibition. However, there are no suitable in vitro methods to distinguish and assess regeneration of severed CNS axons. In this paper we describe the use of a new microfluidic neuron culture device that can be applied quantitatively to investigate the effect of myelin inhibitors on the regeneration of injured CNS axons. The device has two isolated compartments separated by a physical barrier with embedded microgrooves. The device has been successfully used for long-term culture of primary CNS neurons while isolating the somata in one chamber and directing growth of axons to the other chamber. We cultured cortical neurons in the devices and subjected them to reproducible axotomy by vacuum aspiration. After 24 hours, the lengths of regenerated axons were measured for quantitative analysis. NOGO-66 concentrations above 10 nM consistently resulted in ~20% reduction in length of regenerated axons. MAG protein also inhibited axonal regeneration. The length of regenerated axons decreased with addition of myelin inhibitory proteins after injury. These results suggest that the device can serve as an in vitro model for axonal injury and regeneration by simulating the microenvironment around the injury site.
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Hellman, Amy N.,Vahidi, Behrad,Kim, Hyung Joon,Mismar, Wael,Steward, Oswald,Jeon, Noo Li,Venugopalan, Vasan Royal Society of Chemistry 2010 Lab on a chip Vol.10 No.16
<P>We describe the integrated use of pulsed laser microbeam irradiation and microfluidic cell culture methods to examine the dynamics of axonal injury and regeneration <I>in vitro</I>. Microfabrication methods are used to place high purity dissociated central nervous system neurons in specific regions that allow the axons to interact with permissive and inhibitory substrates. Acute injury to neuron bundles is produced <I>via</I> the delivery of single 180 ps duration, λ = 532 nm laser pulses. Laser pulse energies of 400 nJ and 800 nJ produce partial and complete transection of the axons, respectively, resulting in elliptical lesions 25 μm and 50 μm in size. The dynamics of the resulting degeneration and regrowth of proximal and distal axonal segments are examined for up to 8 h using time-lapse microscopy. We find the proximal and distal dieback distances from the site of laser microbeam irradiation to be roughly equal for both partial and complete transection of the axons. In addition, distinct growth cones emerge from the proximal neurite segments within 1–2 h post-injury, followed by a uniform front of regenerating axons that originate from the proximal segment and traverse the injury site within 8 h. We also examine the use of EGTA to chelate the extracellular calcium and potentially reduce the severity of the axonal degeneration following injury. While we find the addition of EGTA to reduce the severity of the initial dieback, it also hampers neurite repair and interferes with the formation of neuronal growth cones to traverse the injury site. This integrated use of laser microbeam dissection within a micropatterned cell culture system to produce precise zones of neuronal injury shows potential for high-throughput screening of agents to promote neuronal regeneration.</P> <P>Graphic Abstract</P><P>We describe the integrated use of pulsed laser microbeam irradiation and microfluidic cell culture to examine the dynamics of axonal injury and regeneration <I>in vitro</I>. <IMG SRC='http://pubs.rsc.org/services/images/RSCpubs.ePlatform.Service.FreeContent.ImageService.svc/ImageService/image/GA?id=b927153h'> </P>
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Integrated microfluidics platforms for investigating injury and regeneration of CNS axons.
Kim, Hyung Joon,Park, Jeong Won,Byun, Jae Hwan,Vahidi, Behrad,Rhee, Seog Woo,Jeon, Noo Li Pergamon Press [etc.] 2012 Annals of biomedical engineering Vol.40 No.6
<P>We describe the development of experimental platforms to quantify the regeneration of injured central nervous system (CNS) neurons by combining engineering technologies and primary neuronal cultures. Although the regeneration of CNS neurons is an important area of research, there are no currently available methods to screen for drugs. Conventional tissue culture based on Petri dish does not provide controlled microenvironment for the neurons and only provide qualitative information. In this review, we introduced the recent advances to generate in vitro model system that is capable of mimicking the niche of CNS injury and regeneration and also of testing candidate drugs. We reconstructed the microenvironment of the regeneration of CNS neurons after injury to provide as in vivo like model system where the soluble and surface bounded inhibitors for regeneration are presented in physiologically relevant manner using microfluidics and surface patterning methods. The ability to control factors and also to monitor them using live cell imaging allowed us to develop quantitative assays that can be used to compare various drug candidates and also to understand the basic mechanism behind nerve regeneration after injury.</P>
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