Quantitative analysis of cell composition and purity of human pancreatic islet preparations.
Quantitative analysis of cell composition and purity of human pancreatic islet preparations.
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DOI：
10.1038/labinvest.2010.124
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发表时间：
2010-11
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尽管人类胰岛移植的结果有所改善，但胰岛制剂的特征仍然不明确。本研究采用光学显微镜（LM）和电子显微镜（EM）来表征33个用于临床移植的胰岛制剂。EM允许准确鉴定和定量细胞类型，测量的细胞数量分数（平均值± SEM）为35.6 ± 2.1% β细胞、12.6 ± 1.0%非β胰岛细胞（总胰岛细胞48.3 ± 2.6%）、22.7 ± 1.5%导管细胞和25.3 ± 1.8%腺泡细胞。胰岛细胞中β细胞占73.6 ± 1.7%。为了与文献进行比较，将细胞数量分数、细胞体积和细胞外体积的估计值合并，以将数量分数数据转换为适用于细胞、胰岛和整个制剂的体积分数。建立了这种转换的数学框架。按体积计，β细胞占总胰岛细胞体积的86.5 ± 1.1%，占完整胰岛（包括细胞外体积）的61.2 ± 0.8%，与胰腺中的胰岛相似。我们估计在一个胰岛当量（150 μm直径球体的体积）中有1560 ± 20个细胞，其中1140 ± 15个是β细胞。为了测试相同组织样品的LM分析是否可以提供胰岛制备物纯度的合理估计，通过点计数形态测量法在可从27种临床制备物获得的薄切片上测量胰岛组织的体积分数。通过LM和EM分析测定的单个制剂的胰岛纯度（胰岛体积分数）与极好的一致性线性相关（R2 = 0.95）。然而，常规双硫腙染色的胰岛纯度明显更高，高估了20-30%。因此，EM和LM都提供了准确的方法来确定人类胰岛制剂的细胞组成，并可以帮助我们了解文献中胰岛组成的许多差异。
Despite improvements in outcomes for human islet transplantation, characterization of islet preparations remains poorly defined. This study used both light (LM) and electron microscopy (EM) to characterize 33 islet preparations used for clinical transplants. EM allowed accurate identification and quantification of cell types with measured cell number fractions (mean ± SEM) 35.6 ± 2.1% β-cells, 12.6 ± 1.0% non-β-islet cells, (48.3 ± 2.6% total islet cells), 22.7 ± 1.5% duct cells, and 25.3 ± 1.8% acinar cells. Of the islet cells, 73.6 ± 1.7% were β cells. For comparison to the literature, estimates of cell number fraction, cell volume, and extracellular volume were combined to convert number fraction data to volume fractions applicable to cells, islets, and the entire preparation. The mathematical framework for this conversion was developed. By volume, β cells were 86.5 ± 1.1% of the total islet cell volume and 61.2 ± 0.8% of intact islets (including the extracellular volume), which is similar to that of islets in the pancreas. Our estimates gave 1560 ± 20 cells in an islet equivalent (volume of 150-μm diameter sphere), of which 1140 ± 15 were β cells. To test if LM analysis of the same tissue samples could provide reasonable estimates of purity of the islet preparations, volume fraction islet tissue was measured on thin sections available from 27 of the clinical preparations by point counting morphometrics. Islet purity (islet volume fraction) of individual preparations determined by LM and EM analysis correlated linearly with excellent agreement (R2 = 0.95). However, islet purity by conventional dithizone staining was substantially higher with a 20-30% overestimation. Thus, both EM and LM provide accurate methods to determine the cell composition of human islets preparations and can help us understand many of the discrepancies of islet composition in the literature.