Department of Radiation Oncology, Stanford University, California, 94305, USA.
Clin Cancer Res. 2011 Dec 1;17(23):7265-72. doi: 10.1158/1078-0432.CCR-11-0179. Epub 2011 Oct 13.
To assess aldehyde dehydrogenase (ALDH) expression in adult human and murine submandibular gland (SMG) stem cells and to determine the effect of ALDH3 activation in SMG stem cell enrichment.
Adult human and murine SMG stem cells were selected by cell surface markers (CD34 for human and c-Kit for mouse) and characterized for various other stem cell surface markers by flow cytometry and ALDH isozymes expression by quantitative reverse transcriptase PCR. Sphere formation and bromodeoxyuridine (BrdUrd) incorporation assays were used on selected cells to confirm their renewal capacity and three-dimensional (3D) collagen matrix culture was applied to observe differentiation. To determine whether ALDH3 activation would increase stem cell yield, adult mice were infused with a novel ALDH3 activator (Alda-89) or with vehicle followed by quantification of c-Kit(+)/CD90(+) SMG stem cells and BrdUrd(+) salispheres.
More than 99% of CD34(+) huSMG stem cells stained positive for c-Kit, CD90 and 70% colocalized with CD44, Nestin. Similarly, 73.8% c-Kit(+) mSMG stem cells colocalized with Sca-1, whereas 80.7% with CD90. Functionally, these cells formed BrdUrd(+) salispheres, which differentiated into acinar- and ductal-like structures when cultured in 3D collagen. Both adult human and murine SMG stem cells showed higher expression of ALDH3 than in their non-stem cells and 84% of these cells have measurable ALDH1 activity. Alda-89 infusion in adult mice significantly increased c-Kit(+)/CD90(+) SMG population and BrdUrd(+) sphere formation compared with control.
This is the first study to characterize expression of different ALDH isozymes in SMG stem cells. In vivo activation of ALDH3 can increase SMG stem cell yield, thus providing a novel means for SMG stem cell enrichment for future stem cell therapy.
评估醛脱氢酶（ALDH）在成人和鼠下颌下腺（SMG）干细胞中的表达，并确定 ALDH3 激活对 SMG 干细胞富集的影响。
通过细胞表面标志物（人 CD34，鼠 c-Kit）选择成人和鼠 SMG 干细胞，并通过流式细胞术和定量逆转录 PCR 检测各种其他干细胞表面标志物，检测 ALDH 同工酶的表达。通过球体形成和溴脱氧尿苷（BrdUrd）掺入试验对选定的细胞进行验证，以确认其更新能力，并应用三维（3D）胶原基质培养观察分化。为了确定 ALDH3 激活是否会增加干细胞产量，将新型 ALDH3 激活剂（Alda-89）或载体输注到成年小鼠中，然后定量分析 c-Kit（+）/CD90（+）SMG 干细胞和 BrdUrd（+）唾液球体。
超过 99%的 CD34（+）huSMG 干细胞对 c-Kit、CD90 和 70%的 CD44、Nestin 呈阳性染色。同样，73.8%的 c-Kit（+）mSMG 干细胞与 Sca-1 共定位，而 80.7%与 CD90 共定位。功能上，这些细胞形成 BrdUrd（+）唾液球体，当在 3D 胶原中培养时，这些球体分化为腺泡样和导管样结构。成人的人和鼠的 SMG 干细胞都表现出比非干细胞更高的 ALDH3 表达，其中 84%的细胞具有可测量的 ALDH1 活性。与对照组相比，Alda-89 输注到成年小鼠中显著增加了 c-Kit（+）/CD90（+）SMG 群体和 BrdUrd（+）球体的形成。
这是首次研究 ALDH 同工酶在 SMG 干细胞中的表达。ALDH3 的体内激活可以增加 SMG 干细胞的产量，从而为 SMG 干细胞的富集提供了一种新的方法，为未来的干细胞治疗提供了可能。
