Isolation, characterization, and in vitro proliferation of canine mesenchymal stem cells derived from bone marrow, adipose tissue, muscle, and periosteum
Isolation, characterization, and in vitro proliferation of canine mesenchymal stem cells derived from bone marrow, adipose tissue, muscle, and periosteum
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DOI：
10.2460/ajvr.73.8.1305
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发表时间：
2012-08-01
影响因子：
1
通讯作者：
Nino-Fong, Rodolfo
中科院分区：
文献类型：
--
作者：
Kisiel, Agatha H.;McDuffee, Laurie A.;Nino-Fong, Rodolfo
目的：从犬肌肉和骨膜中分离和表征间充质干细胞（MSCs），并比较骨髓、脂肪组织、肌肉和骨膜来源的间充质干细胞（BMSCs、AMSCs、MMSCs和PMSCs）的增殖能力。样本-7犬类尸体。程序-根据形态学、mscs相关细胞表面标记物的免疫荧光和多能性相关转录因子的表达来表征mscs。形态学和组织化学方法用于评估在成脂、成骨和软骨培养基中培养的MSCs的分化。碱性磷酸酶、RUNX2、OSTERIX和OSTEOPONTIN的信使核糖核酸表达作为成骨分化的标志。分别在24,48,72和96小时对传代1 MSCs进行计数，以确定组织特异性MSC增殖能力。计算融合传代-1型间充质干细胞每克组织的间充质干细胞产量。结果：BMSCs、AMSCs、MMSCs和PMSCs的成功分离是基于形态学的；CD44和CD90的表达；CD34、CD45不表达；SOX2、OCT4、NANOG的mRNA表达；以及脂肪和成骨分化。在4天的培养期内，BMSCs、AMSCs、MMSCs和PMSCs的增殖能力没有显著差异。在传代1中融合后，骨膜每克组织的MSC产量显著提高（平均+/- SD为19,400,000 +/- 12,800,000 /g骨膜，平均+/- SD为13 +/- 1.64天）。结论与临床意义：结果提示犬的肌肉和骨膜可能是骨髓间充质干细胞的来源。骨膜是间充质干细胞产量的优良组织来源，可能在同种异体应用中有用。（美国兽医杂志2012；73:1305-1317）
Objective-To isolate and characterize mesenchymal stem cells (MSCs) from canine muscle and periosteum and compare proliferative capacities of bone marrow-, adipose tissue-, muscle-, and periosteum-derived MSCs (BMSCs, AMSCs, MMSCs, and PMSCs, respectively).Sample-7 canine cadavers.Procedures-MSCs were characterized on the basis of morphology, immunofluorescence of MSC-associated cell surface markers, and expression of pluripotency-associated transcription factors. Morphological and histochemical methods were used to evaluate differentiation of MSCs cultured in adipogenic, osteogenic, and chondrogenic media. Messenger ribonucleic acid expression of alkaline phosphatase, RUNX2, OSTERIX, and OSTEOPONTIN were evaluated as markers for osteogenic differentiation. Passage-1 MSCs were counted at 24, 48, 72, and 96 hours to determine tissue-specific MSC proliferative capacity. Mesenchymal stem cell yield per gram of tissue was calculated for confluent passage-1 MSCs.Results-Successful isolation of BMSCs, AMSCs, MMSCs, and PMSCs was determined on the basis of morphology; expression of CD44 and CD90; no expression of CD34 and CD45; mRNA expression of SOX2, OCT4, and NANOG; and adipogenic and osteogenic differentiation. Proliferative capacity was not significantly different among BMSCs, AMSCs, MMSCs, and PMSCs over a 4-day culture period. Periosteum provided a significantly higher MSC yield per gram of tissue once confluent in passage 1 (mean +/- SD of 19,400,000 +/- 12,800,000 of PMSCs/g of periosteum obtained in a mean +/- SD of 13 +/- 1.64 days).Conclusions and Clinical Relevance-Results indicated that canine muscle and periosteum may be sources of MSCs. Periosteum was a superior tissue source for MSC yield and may be useful in allogenic applications. (Am J Vet Res 2012;73:1305-1317)