Pulmonary artery hypertension (PAH) is a progressive disease, with an estimated median survival of 2.8 years from the time of diagnosis to death [1]. PAH has a complex pathogenesis and is characterized by arteriolar wall remodeling, elevated pulmonary artery pressure, and right ventricular (RV) hypertrophy [2]. The increased pulmonary vascular resistance and excessive proliferation of endothelial cells in the pulmonary artery lead to pulmonary vascular remodeling [3, 4, 5], with widespread loss of the pulmonary microvasculature carrying blood from the heart to the lungs [6]. Although current treatments may prolong and improve the quality of life of patients, the long-term outcome of this disorder is lacking.

Mesenchymal stem cells (MSC) represent an archetype of multipotent somatic stem cells. MSC could differentiate into cardiomyocytes, or even cells of nonmesodermal derivation, including hepatocytes and neurons [7, 8, 9, 10, 11]. Although MSC were originally isolated from bone marrow [12, 13], similar populations have been reported in other tissues. Human MSC have now been isolated from adipose tissue [14], umbilical cord blood (UCB) [15, 16, 17, 18], and peripheral blood [19, 20]. Recently, Kogler et al. [18] described a subset of MSC derived from human CB which they called "unrestricted somatic stem cells." All of the research demonstrated that variations in conditions had a significant impact on the potential of the populations generated, even though the initial cell material could be phenotypically identical. MSC have been defined by plastic adherent growth and subsequent expansion under specific culture conditions, by a panel of nonspecific surface antigens, and by in vitro and in vivo differentiation potential [21]. However, the lack of common standards and of a precise definition of initial cell preparations remains a major obstacle for the research and applications of MSC. In the present study, MSC isolated from UCB were used to treat PAH rats, and the genome-wide expression profiles were compared. The advantage of DNA microarray analysis is that it can simultaneously evaluate changes in the relative expression of thousands of genes [22, 23, 24]. The purpose of this study was to investigate the changes of gene expression by microarray analysis after the injection of human UCB-MSCs into monocrotaline (MCT)-induced PAH rats.

In this study, it was demonstrated that human UCB-MSCs (3×10⁶/ml/cm²) which were injected via the external jugular vein were engrafted in the lung tissues of MCT-induced PAH rats. Adverse effects were not observed after the transfusion of human UCB-MSCs. We have demonstrated changes of gene expressions after bone marrow cell transfusion in MCT model of PAH in our previous study [27] and confirmed in the present study. Previously, Umar et al. [28], also reported the application of MSCs from donor rats with PAH reduces RV pressure overload, RV dysfunction, and lung pathology in recipient rats with PAH. Several studies have suggested that immune activation and the secretion of cytokines contribute to the pathogenesis of PAH [29, 30, 31, 32]. Although the etiology of PAH remains unknown despite extensive investigation, the incidence of PAH patients continues to increase in many countries [33, 34, 35, 36]. There is no doubt that human UCB-MSCs have therapeutic utility in the treatment of PAH. Infusion of high doses of human UCB-MSCs effectively reduces systemic inflammation and prevents the development of lung lesions in PAH. Several mechanisms may explain the antiinflammatory effects of human UCB-MSCs in this disease [37, 38, 39]. To investigate the mechanisms underlying the therapeutic effects of human UCB-MSCs, the gene expression profiles of lung tissue obtained 4 weeks after the injection of human UCB-MSCs in PAH rats were examined. The advantage of DNA microarray is that it can simultaneously evaluate changes in the relative expression of thousands of genes [22, 23, 24]. To gain further insight into the mechanism of human UCB-MSCs related to immune processes and genetic factors, the difference of gene expression between the groups injected with human MSCs and UCB-MSCs, the M group and the U group, were investigated. In addition, the difference of gene expression levels after human UCB-MSCs therapy was compared to identify potential candidate genes that might link the systemic immune response to the development of PAH disease by examining the gene expression patterns between the M group and the U group in PAH rats. In the present study, many immunologic processes and genetic factors were attributed to the pathogenesis of PAH. Immunologic abnormalities in the M group of PAH rats reflected marked activation of the immune system, leading to increased cytokine production. The data indicated that there were several genes with differential expression in the U group compared to the M group. The change in expression levels of several genes were confirmed after the injection of human UCB-MSCs. These genes were related to inflammatory response, immune response, lipid metabolism, cell adhesion, cell migration, cell differentiation, apoptosis, cell growth, transport, cell proliferation, transcription, and signal transduction.

Previous reports suggested that naive or gene-modified endothelial progenitors from peripheral blood or bone marrow can ameliorate some of the symptoms of MCT-induced pulmonary hypertension [40, 41]. In the investigation of PAH, gene microarrays have been employed in a variety of study designs performed on a diverse array of cell types and animal species. Animal microarray studies have been performed on both whole lung tissue and microdissected pulmonary vasculature of hypoxic and MCT-induced PAH [42, 43]. These studies have employed "hypothesis-building" strategies, helping to focus attention on potentially novel pathologic pathways. However, gene expression has also been utilized as a biomarker, potentially useful in the classification or identification of an individual's risk of disease [44]. Examination of gene expression profiling from the lungs of two genetically engineered mouse models provided important clues into the pathogenesis of transforming growth factor α (TGF-α)-induced pulmonary fibrosis and disruption of the BMPR2 pathway. The work by Hardie et al. [45]. utilized an advanced tetracycline-inducible transgenic induction of TGF-α with lung specificity.

To define of PAH genes, the scleroderma group was subcategorized into mild and severe. Genes that appeared to correlate with severity of disease included ILA, vascular endothelial growth factor, interleukin-β, and matrix metalloproteinase 9. Moreover, confirmation of previously differentially expressed genes by the study of Bull et al. [22] included ADM, IL7R, ZFP36, GLUL, JUND, and BCL6. To create effective and reliable cell-based therapies for pulmonary hypertension, it is important to clearly identify the population or subpopulations of nonhematopoietic stem or progenitor cells that contribute to particular tissues that provide large numbers of reparative cells while maintaining their differentiation capacity and ability to durably engraft in vivo [46]. Despite the injection of human UCB-MSCs into MCT-induced PAH rats, relatively little is yet known about the relationship between the pulmonary hypertension and MSCs. Therefore, in the current study, we was to investigate the changes of gene expression by microarray analysis after the injection of human UCB-MSCs into MCT-induced PAH rats. Many studies estimated that many of the patients do not respond to a single injection of human UCB-MSCs, and the risk of aneurysm formation is higher in the unresponsive group than among rats who defervesce completely after a single injection of human UCB-MSCs. The limitations of the present study were as follows. The sample size was small, and further analysis with larger samples of other independent sets as well as specific samples such as peripheral blood T cells, monocytes/macrophages would be needed to confirm the results of human UCB-MSCs injection.