Mobilization of human CD34+CD133+ and CD34+CD133− stem cells in vivo by consumption of an extract from Aphanizomenon flos-aquae—related to modulation of CXCR4 expression by an L-selectin ligand?

doi:10.1016/j.carrev.2007.03.004

Cardiovascular Revascularization Medicine

Volume 8, Issue 3, July–September 2007, Pages 189-202

https://doi.org/10.1016/j.carrev.2007.03.004 Get rights and content

Abstract

Objective

The goal of this study was to evaluate effects on human stem cells in vitro and in vivo of an extract from the edible cyanobacterium Aphanizomenon flos-aquae (AFA) enriched for a novel ligand for human CD62L (L-selectin).

Experimental approach

Ligands for CD62L provide a mechanism for stem cell mobilization in conjunction with down-regulation of the CXCR4 chemokine receptor for stromal derived factor 1. Affinity immunoprecipitation was used to identify a novel ligand for CD62L from a water extract from AFA. The effects of AFA water extract on CD62L binding and CXCR4 expression was tested in vitro using human bone marrow CD34⁺ cells and the two progenitor cell lines, KG1a and K562. A double-blind randomized crossover study involving 12 healthy subjects evaluated the effects of consumption on stem cell mobilization in vivo.

Results

An AFA extract rich in the CD62L ligand reduced the fucoidan-mediated externalization of the CXCR4 chemokine receptor on bone marrow CD34⁺ cells by 30% and the CD62L⁺ CD34⁺ cell line KG1A by 50% but did not alter the CXCR4 expression levels on the CD34⁻ cell line K562. A transient, 18% increase in numbers of circulating CD34⁺ stem cells maximized 1 hour after consumption (P<.0003). When 3 noncompliant volunteers were removed from analysis, the increase in CD34⁺ cells was 25% (P<.0001).

Conclusion

AFA water extract contains a novel ligand for CD62L. It modulates CXCR4 expression on CD34⁺ bone marrow cells in vitro and triggers the mobilization of CD34⁺ CD133⁺ and CD34⁺ CD133⁻ cells in vivo.

Introduction

Much recent research has focused on the role of selectins and their ligands in mobilization of bone marrow stem cells. L-selectin belongs to the selectin family of cell adhesion molecules involved in cellular migration during normal immunosurveillance and inflammatory conditions. L-selectin is best known as a homing molecule for recirculating lymphocytes to recognize high endothelial venules during the process of extravasation [1], [2], [3] and for leukocytes to recognize and home to inflamed tissues [4], [5], [6], [7], [8]. However, L-selectin plays significant roles in other physiological cell adhesion processes as well, including the retention vs. release of bone marrow stem cells into the blood circulation [9], [10], [11].

Of special importance are findings that engagement of L-selectin by some ligands will modulate the expression of the CXCR4 chemokine receptor [12]. The CXCR4 receptor specifically recognizes the chemokine stromal derived factor 1 (SDF-1), which acts as a potent chemoattractant for stem cells and assists in retaining stem cells within the bone marrow environment [13], [14], [15], [16]. The chemoattractant properties of SDF-1 on stem cells were shown in vitro [17] as well as in vivo to be directly associated with recruitment of stem cells into kidney [18] and liver [19]. The mobilization of recruitment of stem cells is associated with repair of the central nervous system [20], [21], heart [22], [23], and other tissues [24]. Stem cell mobilization and homing involve a series of G-protein-coupled receptors that can interact with each other as well as with adhesion molecules [25], [26]. It is proposed that loss of responsiveness towards CXCR4 may be one of several contributing mechanisms that allow some bone marrow stem cells to detach and leave the bone marrow as part of the mobilization process [27], [28].

The use of selectin ligands has been proposed as a mechanism for stem cell mobilization [29]. Some L-selectin ligands (LSLs), including fucoidan and sulfatide, have a proven effect on stem cell mobilization [29], [30]. The mobilization appears to happen in selectin-dependent and -independent mechanisms in tandem. As an example, the sulfated polysaccharide fucoidan can act as an LSL and up-regulate the chemokine receptor CXCR4, a receptor for SDF-1. However, fucoidan also assists stem cell detachment within the marrow by binding to another adhesion receptor, CD11b, during stem cell mobilization [30].

The objective of this study was to evaluate the effects on human stem cells in vitro and in vivo of an extract from Aphanizomenon flos-aquae (AFA), enriched for a novel ligand for human L-selectin. We report here that a novel compound from the blue-green algae AFA binds to the ligand-binding area of human L-selectin. The effect of this compound was tested in various in vitro assays as well as on stem cell mobilization in humans.

Section snippets

Buffers and media

For cell cultures, freshly isolated human marrow cells, as well as the KG1a and K562 cell lines, were resuspended and cultured in RPMI-1640 with 10% fetal calf serum (Gibco, Grand Island, NY, USA), 1% penicillin and streptomycin, and L-glutamine. For immunostaining, cells were washed, resuspended, and stained in phosphate-buffered saline (PBS) containing 0.02% azide and 1% fetal calf serum or bovine serum albumin.

Cyanobacterial extracts

Dried powder of the freshwater blue-green algae AFA was obtained from Desert Lake

AFA contains a ligand for human L-selectin (CD62L)

Using a cell-free affinity purification system in which paramagnetic Dynabeads were coated with a human L-selectin/IgG1 F_c fusion protein, we captured an L-selectin-binding compound from the AFA water extract (AFA-W). Under reducing conditions, this compound showed a distinct double band. Two proteins had apparent molecular weights at 57 and 54 kDa, respectively (Fig. 1). The native protein is larger and estimated to be of an approximate molecular weight of 160–180 kDa.

Comparing band density to

Discussion

Dietary strategies for supporting stem cell biology represent an emerging field of nutritional medicine. The understanding of the effect of nutrition on stem cells needs to include stem cell viability, proliferation, mobilization, and tissue-specific homing. It has been reported that Spirulina consumption may increase erythropoiesis in a mouse model [33]. In addition, antioxidant-rich blueberry and green tea extracts have been reported to increase stem cell life span and proliferation in vitro

Acknowledgments

We are grateful to Kelly M. Patterson and Amber R. Coaty for technical assistance.

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Mobilization of endogenous bone marrow-derived stem cells in a thioacetamide-induced mouse model of liver fibrosis
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A favourable effect of G-CSF administration on survival and clinical parameters in patients with liver failure has also been reported in other studies (Garg et al., 2012; Xing et al., 2013; Wan et al., 2013; Duan et al., 2013; Salama et al., 2014; Singh et al., 2014). Although the magnitude of mobilization induced by SE is milder compared with that triggered by G-CSF (Jensen et al., 2007), our results provide evidence that SE has similar capacity compared to G-SCF and is able to improve liver function and to remedy the histological changes in a TAA mouse model of liver cirrhosis. Similar capabilities were also demonstrated in different target organs.
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During all three phases the lymphocyte–endothelial interaction leads to cellular signaling events, changing resting circulating cells to active effector cells (González-Amaro and Sánchez-Madrid, 1999; Po et al., 1995). Several nutritional products have been shown to support specific lymphocyte subset homing as part of normal immune surveillance (Jensen et al., 2000; 2007; 2011). The present study was undertaken to evaluate this fraction of bovine colostrum for its effects on the innate anti-viral immune system in vitro and in vivo.
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The increased phagocytic activity and rapid transient changes in NK cell numbers suggest that upon consumption, interaction of CLMWF with immune cells in the gut mucosa triggers immediate events with systemic consequences.
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View all citing articles on Scopus

^☆: This project was supported by the Merle-West Center for Medical Research, a nonprofit research organization, Desert Lake Technologies, a harvester of Aphanizomenon flos-aquae, and NIS Labs, an independent natural products research laboratory.

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Cell TherapyMobilization of human CD34+CD133+ and CD34+CD133− stem cells in vivo by consumption of an extract from Aphanizomenon flos-aquae—related to modulation of CXCR4 expression by an L-selectin ligand?☆

Abstract

Objective

Experimental approach

Results

Conclusion

Introduction

Section snippets

Buffers and media

Cyanobacterial extracts

AFA contains a ligand for human L-selectin (CD62L)

Discussion

Acknowledgments

Resuscitation

Exp Toxicol Pathol

Blood

Exp Hematol

Blood

Kidney Int

Neuroscience

Regul Pept

Blood Cells Mol Dis

Blood

Blood

J Neuroimmunol

Blood

Blood

J Am Coll Cardiol

Am Heart J

A cell-surface molecule involved in organ-specific homing of lymphocytes

Nature

Human antigen-specific memory T cells express the homing receptor (LAM-1) necessary for lymphocyte recirculation

Eur J Immunol

Sulphated endothelial ligands for L-selectin in lymphocyte homing and inflammation

Biochem Soc Trans

Insights into selectin function from knockout mice

Thromb Haemost

Ligands for L-selectin: homing, inflammation, and beyond

Annu Rev Immunol

L-selectin: an emerging player in chemokine function

Microcirculation

Endothelial selectins and vascular cell adhesion molecule-1 promote hematopoietic progenitor homing to bone marrow

Proc Natl Acad Sci U S A

Functional selectin ligands mediating human CD34(+) cell interactions with bone marrow endothelium are enhanced postnatally

J Clin Invest

G-CSF induces stem cell mobilization by decreasing bone marrow SDF-1 and up-regulating CXCR4

Nat Immunol

SDF-1 tells stem cells to mind their P's and Z's

J Clin Invest

Cell Therapy
Mobilization of human CD34⁺CD133⁺ and CD34⁺CD133⁻ stem cells in vivo by consumption of an extract from Aphanizomenon flos-aquae—related to modulation of CXCR4 expression by an L-selectin ligand?☆