Alkaline phosphatase in stem cells

Abstract

Alkaline phosphatase is an enzyme commonly expressed in almost all living organisms. In humans and other mammals, determinations of the expression and activity of alkaline phosphatase have frequently been used for cell determination in developmental studies and/or within clinical trials. Alkaline phosphatase also seems to be one of the key markers in the identification of pluripotent embryonic stem as well as related cells. However, alkaline phosphatases exist in some isoenzymes and isoforms, which have tissue specific expressions and functions. Here, the role of alkaline phosphatase as a stem cell marker is discussed in detail. First, we briefly summarize contemporary knowledge of mammalian alkaline phosphatases in general. Second, we focus on the known facts of its role in and potential significance for the identification of stem cells.

Figure 1

Relative alkaline phosphatase activity (a) and mRNA expression of TNAP ((b) qRT-PCR) in various stem/progenitor cells. Bone marrow-derived MSC (bmMSC) has both high AP activity and TNAP expression, followed by ES cells. Relative expression (qRT-PCR) of common pluripotent (Oct-4 (c) and Nanog (d)) and osteogenic (Osteocalcin, OstC (e) and Osteopontin, OstP (f)) genes. A high level of Oct-4 and Nanog documented the real pluripotent status of ES cells. A higher expression of OstC and OstP mRNA in bmMSC documented their osteogenic properties/lineages specification. (ES: mouse embryonic stem cells; NSC: neural stem/progenitor cells; bmMSC: bone marrow mesenchymal stem cells; aMSC: adipose tissue mesenchymal stem cells). Details of the presented assay may be found in our previous work [9, 77]. Primers and conditions for OstC and OstP were as follows: OstC: 5′-CTTGGGTTCTGACTGGGTGT-3′, 5′-GCCCTCTGCAGGTCATAGAG-3′ (212 bp, 60°C); OstP: 5′-TCACCATTCGGATGAGTCTG-3′, 5′-ACTTGTGGCTCTGATGTTCC-3′ (436 bp, 60°C). Error bars indicate ±SD (n ≥ 3, ^* P < 0.05; ^** P < 0.01, ANOVA post hoc Bonferroni's Multiple Comparison Test).

Figure 2

Retinoic acid increased AP activity (a) and mRNA expression of TNAP ((b) qRT-PCR) in mouse ES cells. On the other hand, retinoic acid downregulated the expression of pluripotent markers Oct-4 (c) and Nanog (d) in the same manner as the spontaneous differentiation of ES cells through leukemia inhibitory factor withdrawal [75]. (ES: mouse embryonic stem cells; dES: spontaneously differentiating ES cells for two days; raES: retinoic acid-treated (0.2 μM) ES cells for two days). Details of the presented assay are as in Figure 1. Error bars indicate ±SD (n = 4, ^* P < 0.05; ^** P < 0.01, ANOVA post hoc Bonferroni's Multiple Comparison Test).