下腔静脉间置移植物在小鼠模型中植入
Summary
为了提高我们的细胞和分子neotissue形成的知识，最近被开发的TEVG小鼠模型。移植物植入在C57BL / 6小鼠肾下腔静脉移植物插入。该模型能达到类似的结果那些在我们的临床调查来实现，但在一个远缩短时间过程。
Abstract
接种骨髓单个核细胞（骨髓细胞）生物可降解支架通常用于重建手术治疗先天性心脏畸形。长期的临床结果显示出优异的通畅率，但是，与狭窄的显著发病率。为了探讨血管neotissue形成的细胞和分子机制，防止狭窄发展组织工程血管移植（TEVGs），我们制定了移植物的小鼠模型大约1毫米的直径。首先，TEVGs被从聚乙醇酸无纺布制成可生物降解的管状支架组装毡网眼涂有ε-己内酯和L-丙交酯共聚物。的支架，然后放置于冷冻干燥机，抽真空24小时，并在干燥器中储存直到细胞接种。第二，骨髓是从供体小鼠中收集和单核细胞中分离出密度梯度离心。第三，约百万细胞接种于支架培养的O / N。最后，将种子支架，然后注入在C57BL / 6小鼠肾下腔静脉移植物插入。植入移植物表现出良好的通畅性（> 90％），无血栓栓塞并发症或动脉瘤形成的证据。这种小鼠模型将有助于我们理解和量化neotissue形成的TEVG的细胞和分子机制。
Introduction
先天性心脏缺陷是影响近8％的活产婴儿在美国的严重情况。约25％的婴儿患有先天性心脏缺陷或2.4每1000活产的，需要侵入性治疗在他们的生活1的第一年。最有效的治疗先天性心脏疾病是重建手术。不幸的是，从使用目前可用的血管导管引起的并发症是术后发病率和死亡率的最显著的原因。
为了解决这个问题，我们开发了第一个组织工程血管移植（TEVGs）的临床应用2。 TEVGs从播种与自体骨髓单个核细胞（BM-跨国公司），并作为植入静脉导管先天性心脏手术的可生物降解聚酯管构成。结果表明，优秀的通畅率在1-3年的随访，但狭窄的发生率显著<s高达> 3,4。很明显，是需要更好地了解血管neotissue形成及相关TEVG狭窄的发展机制。为了更好地了解TEVGs的发展和狭窄发展的机制，一个绵羊模型建立5,6。在这个模型中，TEVGs成功转型为生活器皿并分别在这两个形态和原生矿脉的功能类似。这种使用了大量的动物模型是在提供辅助TEVGs的临床应用重要的临床前信息的一个很好的第一步。然而，血管neotissue形成用大动物模型TEVGs的细胞和分子机制的充分了解是由于由于缺乏物种特异的分子工具的血管细胞表型的分子特征局限性的限制。为了克服这些缺点，TEVGs小鼠模型是由原因的迅速发展在小鼠遗传学和其广泛的molecula的研制Ř表征具有缩短的时间刻度的额外优势。
鼠下腔静脉插入模型忠实扼要重述，发生在大动物和人类的新血管形成的过程，但在短得多的时间过程6-9。这里，对于使用可生物降解的支架小型接枝制造一个详细的协议中，BM-MNC收获并进行隔离，脚手架BM-MNC播种，并在小鼠模型中移植物植入进行了描述。
Protocol
Representative Results
Discussion
TEVG的小鼠模型是研究细胞和分子neotissue形成和狭窄的发展机制的重要工具。引晶的BM-MNC中所示的移植物11的种子细胞的两个组织学和SEM图像。细胞接种效率是利用DNA测定7还示出。利用该模型系统中，我们发现，细胞接种降低TEVG狭窄，这是失败的主模式在我们的人体临床试验3的发展的发生率。将接种的细胞迅速从TEVG消失，这表明它们通过旁分泌机制8,12发挥它们?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
这项工作是支持的，部分由一个津贴从美国国立卫生研究院（RO1 HL098228）到CKB。
Materials
| Name | Company | Catalog Number | Comments |
| polyglycolic acid (PGA) felt | Biomedical Structures | Custome ordered | |
| ɛ-caprolactone and L- lactide copolymer P(LA/CL) |
Gunze Inc. | Custome ordered | |
| Pipet tip, 0.1-10 μl | Fisher Sientific | 02-707-456 | |
| Lyophilizer | Labconco | 7070020 | |
| RPMI medium 1604 | gibco | 11875-093 | |
| Petri dish | BD | 353003 | |
| 24 well plate | Corning | 3526 | |
| 15cc tube | BD | 352096 | |
| Ficoll | Sigma | 10831-100ml | Also called 'Histopaque' |
| DPBS | gibco | 14190-144 | |
| Littauer Bone Cutter 4.5" Straight | Roboz | RS-8480 | For BM harvesting |
| Forceps 4.5" | Roboz | RS-8120 | For BM harvesting |
| Scissors 4.5" | Roboz | RS-5912 | For BM harvesting |
| Microscope | Leica | M80 | |
| C57BL/6J (H-2b), Female | Jackson Laboratories | 664 | 8-12 weeks |
| Ketamine Hydrochloride Injection | Hospira Inc. | NDC 0409-2053 | |
| Xylazine Sterile Solution | Akorn Inc. | NADA# 139-236 | |
| ketoprofen | Fort Dodge Animal Health | NDC 0856-4396-01 | |
| Ibuprofen | PrecisionDose | NDC 68094-494-59 | |
| Heparin Sodium | Sagent Pharmaceticals | NDC 25021-400 | |
| Saline solution (Sterile 0.9% Sodium Chloride) | Hospira Inc. | NDC 0409-0138-22 | |
| 0.9% Sodium Chloride Injection | Hospira Inc. | NDC 0409-4888-10 | |
| Petrolatum Ophthalmic Ointment | Dechra Veterinary Products | NDC 17033-211-38 | |
| Iodine Prep Pads | Triad Disposables, Inc. | NDC 50730-3201-1 | |
| Alcohol Prep Pads | McKesson Corp. | NDC 68599-5805-1 | |
| Cotton tipped applicators | Fisher Sientific | 23-400-118 | |
| Fine Scissor | FST | 14028-10 | |
| Micro-Adson Forcep | FST | 11018-12 | |
| Clamp Applying Forcep | FST | 00072-14 | |
| S&T Vascular Clamp | FST | 00396-01 | |
| Spring Scissors | FST | 15008-08 | |
| Colibri Retractors | FST | 17000-04 | |
| Dumont #5 Forcep | FST | 11251-20 | |
| Dumont #7 – Fine Forceps | FST | 11274-20 | |
| Dumont #5/45 Forceps | FST | 11251-35 | |
| Tish Needle Holder/Forceps | Micrins | MI1540 | |
| Black Polyamide Monofilament Suture, 10-0 | AROSurgical Instruments Corporation | TI638402 | For sutureing the graft |
| Black Polyamide Monofilament Suture, 6-0 | AROSurgical Instruments | SN-1956 | For musculature and skin closure |
| Non-Woven Songes | McKesson Corp. | 94442000 | |
| Absorbable hemostat | Ethicon | 1961 | |
| 1 ml Syringe | BD | 309659 | |
| 3 ml Syringe | BD | 309657 | |
| 10 ml Syringe | BD | 309604 | |
| 18G 1 1/2 in, Needle | BD | 305190 | |
| 25G 1 in., Needle | BD | 305125 | |
| 30G 1 in., Needle | BD | 305106 | |
| Warm Water Recircultor | Gaymar | TP-700 | |
| Warming Pad | Gaymar | TP-22G | |
| Trimmer | Wahl | 9854-500 |
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