Magnetic Resonance Analysis of Connective Tissue and Muscle

在一个特定的项目中，我们试图评估使用磁共振成像（MRI）测量临床适用的组织工程软骨系统中不断变化的硫酸化糖胺聚糖（GAG）含量的可靠性。 使用水凝胶、聚（环氧乙烷）二丙烯酸酯（PEODA）样品包封牛软骨细胞（240万个细胞/样品）。使用MRI钆排除法测定发育中软骨的固定电荷密度（FCD）。在孵育9、16、29、36、43和50天后对样品进行MRI实验。随后通过生化程序分析这些时间点的样本，以将MRI衍生的FCD测量值与组织中的真实GAG含量相关联。还处理了样品的组织切片，以揭示GAG浓度的时间差异。我们发现FCD和GAG含量之间的强相关性（R2 = 0.85）被确定为长达36天。然而，当延长到50天时，相关性显著降低（R2 = 0.54）。由此，我们得出结论，软骨细胞包封的水凝胶构建体的FCD在孵育的前36天期间与真实GAG含量良好相关。这项研究表明，MRI衍生的FCD测量可以可靠地解释在早期阶段的评估（5周）的可注射软骨组织工程系统。 在进一步的工作中，我们试图确定组织工程构建物中细胞的分布。 这很难通过非破坏性手段进行研究，例如植入后所需的手段。然而，用含铁颗粒标记细胞可能被证明是解决这个问题的有用方法，因为这种标记细胞的区域已经被证明可以使用磁共振成像容易地检测到。在这项研究中，我们使用FDA批准的超顺磁性氧化铁（SPIO）剂，菲立磁，结合转染剂，标记和可视化与MRI软骨细胞在两个不同的组织工程结构。通过MRI和组织学确定标记细胞位置之间的对应关系。发现主要软骨基质成分的表型、活力和生产不受SPIO标记过程的影响。我们相信，这种可视化和跟踪软骨细胞的方法可能是有用的，在软骨组织工程治疗的进一步发展。 最后，MR也是用于人体受试者的详细代谢研究的优秀模式。 我们感兴趣的是体重、脂质代谢和健康风险之间的联系。 这是一个很难理解和研究的问题。磁共振波谱（MRS）允许非侵入性调查的脂质代谢。我们扩展了现有的二维MRS技术，允许量化的肌细胞内（IMCL）和肌细胞外（EMCL）的脂质室和他们的不饱和度在人类受试者，并将这些结果与BMI。使用肌肉肌酸（Cr）进行标准化，观察到IMCL/Cr随BMI（每组n=8名受试者）的统计学显著（p < 0.01）增加，值为5.9 ± 1.7（BMI < 25）、10.9 ± 1.82（25 < BMI< 30）和13.1 ± 0.87（BMI > 30）。同样，IMCL不饱和度随BMI显著降低（p < 0.01），分别为1.51 ± 0.08、1.30 ± 0.11和0.90 ± 0.14。我们的结论是，脂质代谢的重要方面，可以与二维MRS进行评估，并提出非侵入性测量的不饱和度可以作为与肥胖相关的脂质代谢缺陷的生物标志物。

In one specific project, we sought to assess the reliability in measurement of evolving sulfated glycosaminoglycan (GAG) content in a clinically applicable tissue engineered cartilage system using magnetic resonance imaging (MRI). Samples of the hydrogel, poly(ethylene oxide) diacrylate (PEODA) were used to encapsulate bovine chondrocytes ( 2.4 million cells/ sample). The fixed charge density (FCD) of the developing cartilage was determined using the MRI gadolinium exclusion method. MRI experiments were performed on samples following 9, 16, 29, 36, 43 and 50 days of incubation. Samples from these timepoints were subsequently analyzed via biochemical procedures in order to correlate the MRI-derived FCD measurements with the true GAG content in the tissue. Histological sections of the samples were also processed to reveal temporal differences in the GAG concentration. We found a strong correlation (R2 = 0.85) between FCD and GAG content was determined up to 36 days. However, when extended to 50 days, the correlation decreased significantly (R2 = 0.54). From this, we conclude that FCD of chondrocyte-encapsulated hydrogel constructs correlate well with true GAG content during the first 36 days of incubation. This study demonstrates that MRI-derived FCD measurements can be reliably interpreted in the early stage evaluation ( 5 weeks) of injectable cartilage tissue engineering systems. In further work, we sought to define the distribution of cells within tissue engineered constructs. This is difficult to study through non-destructive means, such as would be required after implantation. However, cell labeling with iron-containing particles may prove to be a useful approach to this problem, since regions of such labeled cells have been shown to be readily detectable using magnetic resonance imaging. In this study, we used the FDA-approved superparamagnetic iron oxide (SPIO) agent, Feridex, in combination with transfection agents to label and visualize with MRI chondrocytes in two different tissue engineered constructs. Correspondence between labeled cell location as determined by MRI and by histology was established. The phenotype, viability and production of major cartilage matrix constituents were found to be unaffected by the SPIO-labeling process. We believe that this method of visualizing and tracking chondrocytes may be useful in the further development of cartilage tissue engineering therapeutics. Lastly, MR is also an excellent modality for detailed metabolic investigations in human subject. We were interested in the link between body weight, lipid metabolism, and health risks. This is poorly understood and difficult to study. Magnetic resonance spectroscopy (MRS) permits non-invasive investigation of lipid metabolism. We extended existing two-dimensional MRS techniques to permit quantification of intramyocellular (IMCL) and extramyocellular (EMCL) lipid compartments and their degree of unsaturation in human subjects, and correlated these results with BMI. Using muscle creatine (Cr) for normalization, a statistically significant (p < 0.01) increase in IMCL/Cr with BMI (n=8 subjects per group) was observed, with values of 5.9 1.7 (BMI < 25), 10.9 1.82 (25 < BMI< 30) and 13.1 0.87 (BMI > 30). Similarly, the degree of IMCL unsaturation decreased significantly (p < 0.01) with BMI, with respective values of 1.51 0.08, 1.30 0.11, and 0.90 0.14. We conclude that important aspects of lipid metabolism can be evaluated with 2-dimensional MRS and propose that degree of unsaturation measured noninvasively may serve as a biomarker for lipid metabolic defects associated with obesity.