Galectin-3 Enhances Proliferation and Angiogenesis of Endothelial Cells Differentiated from Bone Marrow Mesenchymal Stem Cells

doi:10.1016/j.transproceed.2011.10.050

Transplantation Proceedings

Volume 43, Issue 10, December 2011, Pages 3933-3938

https://doi.org/10.1016/j.transproceed.2011.10.050 Get rights and content

Abstract

Purpose

To observe the effects of galectin-3 on proliferation and angiogenesis of endothelial cells differentiated from bone marrow mesenchymal stem (MSCs).

Methods

Cultured MSCs were isolated from bone marrow of Sprague–Dawley rats and purified by gradient centrifugation with lymphocytes separation medium. Cells of passage 3 were differentiated into endothelial cels by vascular endothelial growth factor and basic fibroblast growth factor. These cells were identified as endothelial cells by immunohistochemistry staining and electronic microscopy after 14 days. The cells were cultivated with the galectin-3 at the concentrations of 0.1, 1, and 5 μg/mL for 24 hours. The proliferation of endothelial cells were measured by 3-(4,5-methylth-iazol-2-yl)-diphenyltetrazolium bromide (MTT) and the cell cycle was investigated by using flow cytometry. The functionality of angiogensis was observed when the cells appeared tube formation in presence of glacetin-3.

Results

The proliferation activity, analyzed by MTT method, in the galectin-3 groups (1 and 5 μg/mL) were 0.3002 ± 0.0159 and 0.3514 ± 0.0133, respectively, which were significantly greater than that in the control group (0.2339 ± 0.0041; P < .05). Flow cytometry detection showed that S phase cells (%) are 29.42 ± 0.45, 34.56 ± 0.82, and 52.58 ± 2.84 in groups of 0.1, 1, and 5 μg/mL, respectively, and G₂M phase cells increased from 4.88 ± 1.12 to 5.26 ± 0.45 with the concentrations of 1 and 5 μg/mL, respectively, which demonstrated significant difference compared with the control group (P < .05). The tubular network formation was lengthened significantly compared with the control group (P < .05).

Conclusion

Galectin-3 can promote the proliferation and angiogenesis of endothelial cells differentiated from bone marrow mesenchymal stem cells.

Section snippets

Antibodies and Reagents

LG-DMEM (Hyclone, San Diego, Calif), trypsin-EDTA solution (Beyotime, Jiangsa, China), fetal bovine serum (FBS; Sijiqing, China), percoll (Pharmacia, Kalamazoo, Mich), galectin-3 (Peprotechm Rocky HIll, NJ) vascular enthelial growth actor (VEGF; Peprotech), basic fibroblast growth factor (bFGF; Peprotech), polyclonal ntibody (Promega, Madiso, Wisc), rabbit–anti-mouse polyclonal antibodies against Von Willebrand factor (vWF; Promega), MTT (Amresco, Solon, Oh), propidine iodide (PI, Sigma, Louis,

Cell Culture

Cells were adhered in plastic flasks, P₀ (passage zero) cells appeared in flasks after 2 days of plating with a fibroblast-like, spindle-shaped or polygonal morphology. These cells began to proliferate at about day 4 and gradually grew to form small colonies (Fig 1A). By day 7–8, the number of cellular colonies of different sizes had obviously increased. After approximately 12 days, cells became confluent (Fig 1B). The behavior of passaged MSC was similar to that in primary culture, 0 in the

Discussion

MSCs have shown all criteria of true stem cells, including self-renewal, multilineage differential, and reconstitution of tissue.¹⁰ The use of autologous vascular endothelial progenitor cells seems attractive to the development of engineered vessels as well as to the vascularization of engineered tissues, and may be useful for augment vessel growth in ishcemic tissue.11, 12 MSCs have potency of oriented differentiation into vessel endothelial cells under certain conditions and can be used in

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Cited by (33)

Histochemical characteristics of regressing vessels in the hyaloid vascular system of neonatal mice: Novel implication for vascular atrophy
2018, Experimental Eye Research
Citation Excerpt :
In contrast to their inhibiting the blood supply, there is another possibility that galectin-3-expressing cells accumulate in the hyaloid vessels under ischemic conditions for neovasculogenesis. Galectin-3 induces the differentiation of endothelial cells in vitro and stimulates angiogenesis in vivo (Nangia-Makker et al., 2000; Wan et al., 2011). The angiogenetic effects of galectin-3 contradict our conclusion showing the involvement of galectin-3 in vascular atrophy.
The hyaloid vasculature constitutes a transitory system nourishing the internal structures of the developing eye, but the mechanism of vascular regression and its cell biological characteristics are not fully understood. The present study aimed to reveal the specificity of the hyaloid vessels by a systematic immunohistochemical approach for marker substances of myeloid cells and the extracellular matrix (ECM) in neonatal mice. Macrophages immunoreactive for F4/80, cathepsin D, and LYVE-1 gathered around the vasa hyaloidea propria (VHP), while small round cells in vascular lumen of VHP were selectively immunoreactive for galectin-3; their segmented nuclei and immunoreactivities for Ly-6G, CD11b, and myeloperoxidase indicated their neutrophilic origin. VHP possessed thick ECM and a dense pericyte envelope as demonstrated by immunostaining for laminin, type IV collagen, integrin β1, and NG2. The galectin-3⁺ cells loosely aggregated with numerous erythrocytes in the lumen of hyaloid vessels in a manner reminiscent of vascular congestion. Galectin-3 is known to polymerize and form a complex with ECM and NG2 as well as recruit leukocytes on the endothelium. Observation of galectin-3 KO mice implicated the involvement of galectin-3 in the regression of hyaloid vasculature. Since macrophages may play central roles including blocking of the blood flow and the induction of apoptosis in the regression, galectin-3⁺ neutrophils may play a supportive role in the macrophage-mediated involution of the hyaloid vascular system.
The emerging role of galectins in cardiovascular disease
2016, Vascular Pharmacology
Citation Excerpt :
Wan et al. investigated the effect of galectin-3 on endothelial cells differentiated from rat bone marrow mesenchymal stem cells. They found that galectin-3 enhances endothelial cell proliferation and tube formation, associated with angiogenesis [41]. Pro-angiogenic effects of galectin-3 can also be mediated by an indirect effect via circulating platelets.
Galectins are an ancient family of β-galactoside-specific lectins and consist of 15 different types, each with a specific function. They play a role in the immune system, inflammation, wound healing and carcinogenesis. In particular the role of galectin in cancer is widely studied. Lately, the role of galectins in the development of cardiovascular disease has gained attention. Worldwide cardiovascular disease is still the leading cause of death. In ischemic heart disease, atherosclerosis limits adequate blood flow. Angiogenesis and arteriogenesis are highly important mechanisms relieving ischemia by restoring perfusion to the post-stenotic myocardial area. Galectins act ambiguous, both relieving ischemia and accelerating atherosclerosis. Atherosclerosis can ultimately lead to myocardial infarction or ischemic stroke, which are both associated with galectins. There is also a role for galectins in the development of myocarditis by their influence on inflammatory processes. Moreover, galectin acts as a biomarker for the severity of myocardial ischemia and heart failure.
This review summarizes the association between galectins and the development of multiple cardiovascular diseases such as myocarditis, ischemic stroke, myocardial infarction, heart failure and atrial fibrillation. Furthermore it focuses on the association between galectin and more general mechanisms such as angiogenesis, arteriogenesis and atherosclerosis.
Galectins in angiogenesis: Consequences for gestation
2015, Journal of Reproductive Immunology
Citation Excerpt :
Thus, a possibility that awaits further investigation is the involvement of this lectin in the modulation of trophoblast differentiation during placentation, particularly in view of recent studies showing a significant up-regulation of gal-3 protein upon hypoxia-induced syncytialisation in the BeWo cell line (Hu et al., 2007). Additionally, several studies have attributed pro-angiogenic properties to this lectin (Nangia-Makker et al., 2000b; Fukushi et al., 2004; Wan et al., 2011; Machado et al., 2014), which are anticipated to be significant for vascular responses associated with pregnancy and are discussed in detail in the following sections. As for tandem repeat galectins, the only members identified to date at the foetal–maternal interface include gal-8, gal-9 and gal-4, although the latter has only been described in rat placenta and scant information on its physiological role is available (Arikawa et al., 2012).
Members of the galectin family have been shown to exert several roles in the context of reproduction. They contribute to placentation, maternal immune regulation and facilitate angiogenesis encompassing decidualisation and placenta formation during pregnancy. In the context of neo-vascularisation, galectins have been shown to augment signalling pathways that lead to endothelial cell activation, cell proliferation, migration and tube formation in vitro in addition to angiogenesis in vivo. Angiogenesis during gestation ensures not only proper foetal growth and development, but also maternal health. Consequently, restriction of placental blood flow has major consequences for both foetus and mother, leading to pregnancy diseases. In this review we summarise both the established and the emerging roles of galectin in angiogenesis and discuss the possible implications during healthy and pathological gestation.
The impact of galectin-3 inhibition on aldosterone-induced cardiac and renal injuries
2015, JACC: Heart Failure
This study investigated whether galectin (Gal)-3 inhibition could block aldosterone-induced cardiac and renal fibrosis and improve cardiorenal dysfunction.
Aldosterone is involved in cardiac and renal fibrosis that is associated with the development of cardiorenal injury. However, the mechanisms of these interactions remain unclear. Gal-3, a β-galactoside–binding lectin, is increased in heart failure and kidney injury.
Rats were treated with aldosterone-salt combined with spironolactone (a mineralocorticoid receptor antagonist) or modified citrus pectin (a Gal-3 inhibitor), for 3 weeks. Wild-type and Gal-3 knockout mice were treated with aldosterone for 3 weeks. Hemodynamic, cardiac, and renal parameters were analyzed.
Hypertensive aldosterone-salt–treated rats presented cardiac and renal hypertrophy (at morphometric, cellular, and molecular levels) and dysfunction. Cardiac and renal expressions of Gal-3 as well as levels of molecular markers attesting fibrosis were also augmented by aldosterone-salt treatment. Spironolactone or modified citrus pectin treatment reversed all of these effects. In wild-type mice, aldosterone did not alter blood pressure levels but increased cardiac and renal Gal-3 expression, fibrosis, and renal epithelial-mesenchymal transition. Gal-3 knockout mice were resistant to aldosterone effects.
In experimental hyperaldosteronism, the increase in Gal-3 expression was associated with cardiac and renal fibrosis and dysfunction but was prevented by pharmacological inhibition (modified citrus pectin) or genetic disruption of Gal-3. These data suggest a key role for Gal-3 in cardiorenal remodeling and dysfunction induced by aldosterone. Gal-3 could be used as a new biotarget for specific pharmacological interventions.
Blocked angiogenesis in Galectin-3 null mice does not alter cellular and behavioral recovery after middle cerebral artery occlusion stroke
2014, Neurobiology of Disease
Citation Excerpt :
Gal-3 exacerbates inflammation and striatal damage in neonatal hypoxia/ischemia (Doverhag et al., 2010). Other studies show that it stimulates endothelial cell proliferation and capillary vessel morphogenesis (Nangia-Makker et al., 2000; Wan et al., 2011). Gal-3 is generally only expressed in the central nervous system after brain injury (Mok et al., 2007; Walther et al., 2000), however we found it in the healthy SVZ stem cell niche (Comte et al., 2011).
Angiogenesis is thought to decrease stroke size and improve behavioral outcomes and therefore several clinical trials are seeking to augment it. Galectin-3 (Gal-3) expression increases after middle cerebral artery occlusion (MCAO) and has been proposed to limit damage 3 days after stroke. We carried out mild MCAO that damages the striatum but spares the cerebral cortex and SVZ. Gal-3 gene deletion prevented vascular endothelial growth factor (VEGF) upregulation after MCAO. This inhibited post-MCAO increases in endothelial proliferation and angiogenesis in the striatum allowing us to uniquely address the function of angiogenesis in this model of stroke. Apoptosis and infarct size were unchanged in Gal-3^−/− mice 7 and 14 days after MCAO, suggesting that angiogenesis does not affect lesion size. Microglial and astrocyte activation/proliferation after MCAO was similar in wild type and Gal-3^−/− mice. In addition, openfield activity, motor hemiparesis, proprioception, reflex, tremors and grooming behaviors were essentially identical between WT and Gal-3^−/− mice at 1, 3, 7, 10 and 14 days after MCAO, suggesting that penumbral angiogenesis has limited impact on behavioral recovery. In addition to angiogenesis, increased adult subventricular zone (SVZ) neurogenesis is thought to provide neuroprotection after stroke in animal models. SVZ neurogenesis and migration to lesion were overall unaffected by the loss of Gal-3, suggesting no compensation for the lack of angiogenesis in Gal-3^−/− mice. Because angiogenesis and neurogenesis are usually coordinately regulated, identifying their individual effects on stroke has hitherto been difficult. These results show that Gal-3 is necessary for angiogenesis in stroke in a VEGF-dependant manner, but suggest that angiogenesis may be dispensable for post-stroke endogenous repair, therefore drawing into question the clinical utility of augmenting angiogenesis.
Vascular galectins: Regulators of tumor progression and targets for cancer therapy
2013, Cytokine and Growth Factor Reviews
Citation Excerpt :
These effects appear to be concentration dependent as higher concentrations can reduce endothelial function [26] and even induce apoptosis [71]. Angiostimulatory activities have also been attributed to galectin-3 [69,72–76]. Markowska and co-workers showed that galectin-3 induces a concentration dependent angiogenic response in vivo using the mouse corneal micropocket assay [73] while Nangia-Makker et al. showed that galectin-3 acts as a chemo-attractant for endothelial cells [74,77] thereby stimulating in vivo vascularization [74].
Galectins are a family of carbohydrate binding proteins with a broad range of cytokine and growth factor-like functions in multiple steps of cancer progression. They contribute to tumor cell transformation, promote tumor angiogenesis, hamper the anti-tumor immune response, and facilitate tumor metastasis. Consequently, galectins are considered as multifunctional targets for cancer therapy. Interestingly, many of the functions related to tumor progression can be linked to galectins expressed by endothelial cells in the tumor vascular bed. Since the tumor vasculature is an easily accessible target for cancer therapy, understanding how galectins in the tumor endothelium influence cancer progression is important for the translational development of galectin-targeting therapies.

View all citing articles on Scopus

: S.Y. Wan and T.F. Zhang contributed equally to this work.

: Supported in part by the Central Research Laboratory of the First Affiliated of Anhui Medical University and Department of Health of Anhui Province fund.

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Bone marrow and stem cell transplantationGalectin-3 Enhances Proliferation and Angiogenesis of Endothelial Cells Differentiated from Bone Marrow Mesenchymal Stem Cells

Abstract

Purpose

Methods

Results

Conclusion

Section snippets

Antibodies and Reagents

Cell Culture

Discussion

Lancet

Am J Pathol

Allergol Int

Trends Cell Biol

Biochem Biophys Res Commun

Trends Cell Biol

J Surg Res

Am J Pathol

J Biol Chem

J Biol Chem

J Biol Chem

Bone marrow and stem cell transplantation
Galectin-3 Enhances Proliferation and Angiogenesis of Endothelial Cells Differentiated from Bone Marrow Mesenchymal Stem Cells