Cytoplasmic-targeted parvalbumin blocks the proliferation of multipotent mesenchymal stromal cells in prophase

Stem Cell Research & Therapy

Dawidson Assis Gomes¹  Michele A Rodrigues¹  Alfredo M Goes¹  Juliana Lott Carvalho¹  Carolina de Andrade¹  Carolina Soares Barros Melo¹ 

[1] Department of Biochemistry and Immunology, Universidade Federal de Minas Gerais, Av. Antonio Carlos 6627, Belo Horizonte-MG ZIP Code: 31270-901, Brazil

关键词: Cyclins;    Proliferation;    Calcium signaling;    Targeted parvalbumin;    Mesenchymal stem cells;   

Others  :  847138
DOI  :  10.1186/scrt291

 received in 2012-12-10, accepted in 2013-08-02,  发布年份 2013

【 摘 要 】

Introduction

Multipotent mesenchymal stromal cells (MSCs) have gained considerable interest because of their potential use in the treatment of a variety of diseases and injuries. Although remarkable advancements have been made in clinical studies, substantial concerns still regard the safety of MSCs. Some evidence suggests that MSCs can spontaneously generate a population of cells with tumorigenic potential. Thus, studying the molecular mechanisms that control the proliferation of MSCs may be a necessary step toward the development of strategies for safe clinical practice. Ca²⁺ is a second messenger that mediates a wide range of cellular responses, including the regulation of cell proliferation, but little is known about its function in MSCs. The aim of this study was to investigate the effects of targeted Ca²⁺ buffering on MSCs proliferation in vitro.

Methods

Here, we used an adenoviral (Ad) vector encoding the Ca²⁺ chelator protein parvalbumin (PV) fused to a nuclear exclusion signal (NES) and the Discosoma red fluorescent protein (DsRed) to investigate the function of cytoplasmic Ca²⁺ signals on MSC proliferation. Confocal microscopy was used to demonstrate that PV-NES-DsRed was expressed in the cytoplasm. Ca²⁺ signaling was monitored by using Fluo-4-AM. Fluorescence-activated cell sorting (FACS) analysis of cells that were stained with propidium iodide was used as a quantitative measure of cell death. The mitotic index was assessed by immunofluorescence, and the expression of cyclins was examined with Western blot.

Results

Our results show that the Ad-PV-NES-DsRed fusion protein decreased serum-induced Ca²⁺ signaling and blocked the proliferation of rat adipose-derived MSCs (AT-MSCs) in prophase. FACS analysis revealed that Ad-PV-NES-DsRed did not induce cell death in AT-MSCs. Furthermore, Western blot analysis demonstrated that Ad-PV-NES-DsRed reduced extracellular signal-regulated kinase (Erk1/2) phosphorylation and cyclin B1 expression. Buffering cytosolic Ca²⁺ did not alter the expression of cyclins A/D1/D2/D3/E and E2.

Conclusions

Our results show that cytoplasmic Ca²⁺ signals are important for AT-MSCs progression beyond prophase because of their effects on Erk phosphorylation and cyclin B1 expression.

【 授权许可】

   
2013 Melo et al.; licensee BioMed Central Ltd.

【 图 表 】

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