Quantitative analysis by flow cytometry of interstitial cells of Cajal, pacemakers, and mediators of neurotransmission in the gastrointestinal tract
Quantitative analysis by flow cytometry of interstitial cells of Cajal, pacemakers, and mediators of neurotransmission in the gastrointestinal tract
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DOI：
10.1002/cyto.a.20078
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发表时间：
2004-12-01
期刊：
影响因子：
3.7
通讯作者：
Sanders, KM
中科院分区：
文献类型：
--
作者：
Ördög, T;Redelman, D;Sanders, KM
背景：Cajal间质细胞(InterstitialCells of Cajal，ICCs)是一种间质细胞，作为电起搏器和神经肌肉神经传递的介体，在胃肠道运动中发挥重要作用。尽管一些胃肠道运动障碍与ICCs的耗竭有关，但由于其形态多样、网络密度区域性变化以及总体稀缺性，对这些细胞的量化一直是困难的。我们的目的是评价流式细胞术(FCM)在ICCs计数中的应用。方法：我们用荧光试剂盒(CD117)抗体鉴定活的胃肠道肌肉或原代培养的有或不有干细胞因子(SCF)表达的STO成纤维细胞中的ICCs。因为这项技术也非特异性地标记常驻巨噬细胞，我们用额外的荧光抗体鉴定了后者。结果：18只小鼠远端胃内ICCs占细胞总数的1.63+/-0.17%，近端结肠占5.85+/-0.84%，远端结肠占6.28+/-0.61%(n=3)。在几乎缺乏ICCs的W/W-V小鼠(n=5)的胃底肌中，检测到非常少的Kit(+)细胞。在表达膜结合型SCF的STO成纤维细胞(n=6)上培养的肠上皮细胞中，FCM检测到的Kit+ICCs数是可溶性SCF刺激的细胞(n=6)的2.6~7.3倍。结论：FCM是一种灵敏、特异的ICCs定量方法。(C)2004年Wiley-Liss公司
Background: Interstitial cells of Cajal (ICCs) are mesenchymal cells that play critical roles in gastrointestinal motility as electrical pacemakers and mediators of neuromuscular neurotransmission. Although depletions of ICCs have been implicated in several gastrointestinal motor disorders, quantification of these cells has been difficult due to their varied morphology, regionally changing network density, and overall scarcity. Our goal was to evaluate flow cytometry (FCM) for the enumeration of ICCs.Methods: We identified murine ICCs in live gastrointestinal muscles or primary cell cultures grown in the presence or absence of stem cell factor (SCF)-expressing STO fibroblasts with fluorescent Kit (CD117) antibodies. Because this technique also labels resident macrophages nonspecifically, we identified the latter with additional fluorescent antibodies. Dispersed cells were analyzed by FCM.Results: ICCs represented 1.63 +/- 0.17% of the total cell count in the distal stomach (n = 18 mice) and 5.85 +/- 0.84% in the proximal colon and 6.28 +/- 0.61% in the distal colon (n = 3 mice). In fundic muscles of W/W-V mice (n = 5) that virtually lack ICCs, very few Kit(+) cells were detected. FCM identified approximately 2.6- to 7.3-fold more Kit+ ICCs in small intestinal cell cultures grown on STO fibroblasts expressing membrane-bound SCF (n = 6) than in cultures stimulated with soluble SCF (n = 6).Conclusions: FCM is a sensitive and specific method for the unbiased quantification of ICCs. (C) 2004 Wiley-Liss, Inc.