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ABSTRACT: Innate immune system has been known to play an important role in inhibiting the malignant transformation, tumor progression, and invasion. However, the mechanistic basis remains ambiguous. Despite polyclonality of human γδ T cells, Vγ2Vδ2 T cell subset was shown to recognize and limit the growth of various tumors at various degrees. The differential recognition of the tumor cells by Vγ2Vδ2 T cells are yet to be defined. Our current study reveals that γδ T cells limit in vitro growth of most breast tumor cells, such as SkBr7 (HER2+), MCF7 (ER+) and MDA-MB-231 (ER-) by inhibiting their survival and inducing apoptosis, except BrCa-MZ01 (PR+) cells. To investigate detail mechanisms of anti-neoplastic effects, we found that cell death was associated with the surface expression levels of MICA/B and ICAM1. Molecular signaling analysis demonstrated that inhibition of cell growth by γδ T cells was associated with the lower expression levels of cell survival-related molecules such as AKT, ERK and concomitant upregulation of apoptosis-related molecules, such as PARP, cleaved caspase 3, and tumor suppressor genes PTEN and P53. However, opposite molecular signaling was observed in the resistant cell line after co-culture with γδ T cells. In vivo, anti-neoplastic effects of γδ T cells were also documented, where tumor growth was inhibited due to the downregulation of survival signals, strong induction of apoptotic molecules, disruption of microvasculature, and increased infiltration of tumor associated macrophages (TAMs). These findings reveal that a complex molecular signaling is involved in γδ T cell-mediated anti-neoplastic effects. © 2013 Wiley Periodicals, Inc.
International Journal of Cancer 04/2013; · 5.44 Impact Factor
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Reeva Aggarwal,
Jingwei Lu,
Suman Kanji,
Matthew Joseph,
Manjusri Das,
Garrett J Noble,
Brooke K McMichael,
Sudha Agarwal,
Richard T Hart,
Zongyang Sun,
Beth S Lee,
Thomas J Rosol,
Rebecca Jackson,
Hai-Quan Mao, Vincent J Pompili,
Hiranmoy Das
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ABSTRACT: Osteoporosis is a bone disorder associated with loss of bone mineral density and micro architecture. A balance of osteoblasts and osteoclasts activities maintains bone homeostasis. Increased bone loss due to increased osteoclast and decreased osteoblast activities is considered as an underlying cause of osteoporosis.
The cures for osteoporosis are limited, consequently the potential of CD34+ cell therapies is currently being considered. We developed a nanofiber-based expansion technology to obtain adequate numbers of CD34(+) cells isolated from human umbilical cord blood, for therapeutic applications. Herein, we show that CD34(+) cells could be differentiated into osteoblastic lineage, in vitro. Systemically delivered CD34(+) cells home to the bone marrow and significantly improve bone deposition, bone mineral density and bone micro-architecture in osteoporotic mice. The elevated levels of osteocalcin, IL-10, GM-CSF, and decreased levels of MCP-1 in serum parallel the improvements in bone micro-architecture. Furthermore, CD34(+) cells improved osteoblast activity and concurrently impaired osteoclast differentiation, maturation and functionality.
These findings demonstrate a novel approach utilizing nanofiber-expanded CD34(+) cells as a therapeutic application for the treatment of osteoporosis.
PLoS ONE 01/2012; 7(6):e39365. · 4.09 Impact Factor
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ABSTRACT: Ischemia related diseases are on rise worldwide and have been shown to cause irreversible damage to the cells due to the blockage
of blood supply to the tissue. Conventional therapies are less effective as they do not consider repair of the damaged tissues.
Thus, alternative, stem cell-based therapies are currently under investigation. For example, hematopoietic stem cells (HSCs)
were shown to give rise to vascular cells involved in neoangiogenesis; so, they have been tested in variety of animal models
and small-scale clinical trials. Improvement in blood flow and tissue functionality was observed and adverse effects were
not apparent. However, success of stem cell therapy is limited by the number of functional stem cells for clinical application.
Numerous attempts are underway to address this issue via strategies that involve ex vivo expansion of stem cells preserving
their stemness. This chapter outlines the mechanism of therapeutic angiogenesis, sources of HSCs, various methods of ex vivo
expansion of HSCs via genetic regulators, cytokines and biomaterial scaffolds, and their preclinical and clinical applications.
KeywordsHematopoietic stem cells-Ischemia-Angiogenic-Vasculogenesis-Stromal cells-Thrombopoietin
12/2011: pages 219-229;
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ABSTRACT: Vasculogenesis and angiogenesis are the major forms of blood vessel formation. Angiogenesis is the process where new vessels grow from pre-existing blood vessels, and is very important in the functional recovery of pathological conditions, such as wound healing and ischemic heart diseases. The development of better animal model and imaging technologies in past decades has greatly enriched our understanding on vasculogenesis and angiogenesis processes. Hypoxia turned out to be an important driving force for angiogenesis in various ischemic conditions. It stimulates expression of many growth factors like vascular endothelial growth factor, platelet-derived growth factor, insulin-like growth factor, and fibroblast growth factor, which play critical role in induction of angiogenesis. Other cellular components like monocytes, T cells, neutrophils, and platelets also play significant role in induction and regulation of angiogenesis. Various stem/progenitor cells also being recruited to the ischemic sites play crucial role in the angiogenesis process. Pre-clinical studies showed that stem/progenitor cells with/without combination of growth factors induce neovascularization in the ischemic tissues in various animal models. In this review, we will discuss about the fundamental factors that regulate the angiogenesis process and the use of stem cells as therapeutic regime for the treatment of ischemic diseases.
Cell biochemistry and biophysics 10/2011; · 3.34 Impact Factor
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ABSTRACT: Maintenance of ex vivo hematopoietic stem cells (HSC) pool and its differentiated progeny is regulated by complex network of transcriptional factors, cell cycle proteins, extracellular matrix, and their microenvironment through an orchestrated fashion. Strides have been made to understand the mechanisms regulating in vivo quiescence and proliferation of HSCs to develop strategies for ex vivo expansion. Ex vivo expansion of HSCs is important to procure sufficient number of stem cells and as easily available source for HSC transplants for patients suffering from hematological disorders and malignancies. Our lab has established a nanofiber-based ex vivo expansion strategy for HSCs, while preserving their stem cell characteristics. Ex vivo expanded cells were also found biologically functional in various disease models. However, the therapeutic potential of expanded stem cells at clinical level still needs to be verified. This review outlines transcriptional factors that regulate development of HSCs and their commitment, genes that regulate cell cycle status, studies that attempt to develop an effective and efficient protocol for ex vivo expansion of HSCs and application of HSC in various non-malignant and malignant disorders. Overall the goal of the current review is to deliver an understanding of factors that are critical in resolving the challenges that limit the expansion of HSCs in vivo and ex vivo.
Current Molecular Medicine 10/2011; 12(1):34-49. · 5.10 Impact Factor
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ABSTRACT: Maintenance of hematopoietic stem cells (HSCs) pool depends on fine balance between self-renewal and differentiation of HSCs. HSCs normally reside within the bone marrow niche of an adult mammal. The embryonic development of HSCs is a complex process that involves the migration of developing HSCs in multiple anatomical sites. Throughout the process, developing HSCs receive internal (transcriptional program) and external (HSC niche) signals, which direct them to maintain balance between self-renewal and differentiation, also to generate a pool of HSCs. In physiological condition HSCs differentiate into all mature cell types present in the blood. However, in pathological condition they may differentiate into non-hematological cells according to the need of the body. It was shown that HSCs can transdifferentiate into cell types that do not belong to the hematopoietic system suggests a complete paradigm shift of the hierarchical hematopoietic tree. This review describes the developmental origins and regulation of HSCs focusing on developmental signals that induce the adult hematopoietic stem cell program, as these informations are very critical for manipulating conditions for expansion of HSCs in ex vivo condition. This review also states clinical application and related patents using HSC.
Recent patents on biotechnology. 04/2011; 5(1):40-53.
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ABSTRACT: The Phase I clinical study was designed to assess the safety and feasibility of a dose escalating intracoronary infusion of autologous bone marrow (BM)-derived CD133+ stem cell therapy to the patients with chronic total occlusion (CTO) and ischemia. Nine patients were received CD133+ cells into epicardial vessels supplying collateral flow to areas of viable ischemic myocardium in the distribution of the CTO. There were no major adverse cardiac events (MACE), revascularization, re-admission to the hospital secondary to angina, or acute myocardial infarction (AMI) for the 24-month period following cellular infusion. In addition, there were no periprocedural infusion-related complications including malignant arrhythmias, loss of normal coronary blood flow or acute neurologic events. Cardiac enzymes were negative in all patients. There was an improvement in the degree of ischemic myocardium, which was accompanied by a trend towards reduction in anginal symptoms. Intracoronary infusion of autologous CD133+ marrow-derived cells is safe and feasible. Cellular therapy with CD133+ cells to reduce anginal symptoms and to improve ischemia in patients with CTO awaits clinical investigation in Phase II/III trials.
Frontiers in bioscience (Elite edition) 01/2011; 3:506-14.
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Journal of the American College of Cardiology 06/2010; 55(22):2510. · 14.16 Impact Factor
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ABSTRACT: Hematopoietic stem cell transplantation has been applied as a standard procedure of treatment for hematological disorders like multiple myeloma and leukemia for several decades. Various sources of stem cells like bone marrow, peripheral blood and umbilical cord blood are used for the transplantation. Among these umbilical cord blood is currently preferred due to the primitiveness of the derived stem cells and minimal possibilities of graft versus host disease or development of graft induced tumors. One of the problems for these sources is the procurement of sufficient number of donor stem cells. Inadequate number of cells may lead to delayed recovery and decrease survivability of the patient. Thus to overcome the limitation of stem cell number, development of an ex-vivo expansion technology is critically important. The recent emerging technology using nanofiber in combination with growth factors has made a significant improvement to the field of regenerative medicine and a couple of patents have been filed. In this review, we will focus on factors regulating hematopoietic stem cell self-renewal and expansion emphasizing on nanofiber as a supporting matrix.
Recent patents on nanotechnology. 06/2010; 4(2):125-35.
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ABSTRACT: Stem cell therapy is currently considered as an important regime for repairing, replacing or enhancing the biological functions of the damaged tissues. Among adult stem cells, hematopoietic stem cells (HSCs) are commonly used for cure of hematological disorders. However, the number of HSCs obtained from sources like bone marrow, peripheral or umbilical cord blood is not sufficient for routine clinical application. Thus, ex-vivo expansion of HSCs becomes critically important. Ex-vivo culture and expansion of stem cells are challenging, as stem cells differentiate in culture rather than self-renew. Lack of clarity about the factors responsible for quiescence and differentiation of HSCs, investigators struggled to optimize conditions for ex vivo expansion. As we understand better, various strategies can be incorporated to mimic in vivo conditions for successful expansion of stem cells. However, characterization and biological functionality should also be tested for expanded stem cells prior to clinical application. To treat ischemia by enhancing therapeutic angiogenesis and neo-vascularization, the role of genetic modification of HSCs with pro-angiogenic factors is the focus of this review.
Frontiers in Bioscience 01/2010; 15:854-71. · 3.52 Impact Factor
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Marcie R Finney,
Laura R Fanning,
Matthew E Joseph,
Jonathan L Goldberg,
Nicholas J Greco,
Shyam Bhakta,
Daniel G Winter,
Margaret Forster,
Paul E Scheid,
Marwa Sabe, Vincent J Pompili,
Mary J Laughlin
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ABSTRACT: Current clinical trials utilize non-selected bone marrow (BM) mononuclear cells (MNC) to augment vasculo genesis within ischemic vascular beds. Recent reports have identified a diminished number and function of hemat-opoietic stem cells (HSC) from aged and diseased patients. Umbilical cord blood (UCB) provides a potential robust allo-geneic source of HSC for therapeutic vasculogenesis.
MNC and magnetically isolated CD133(+) cells were assessed for viability (trypan blue) and surface phenotype (flow cytometry). To test in vivo functionality of the cells, NOD/SCID mice underwent ligation of the right femoral artery followed immediately by cell injection. Blood flow recovery, necrosis, BM engraftment of human cells and histologic capillary density were determined. Cells were tested for potential mechanisms mediating the in vivo effects, including migration, cytokine secretion and angiogenic augmentation (Matrigel assays).
Surface expression analysis showed CD31 (PECAM) expression was greatly increased in UCB CD133(+) cells compared with BM MNC. At 28 days, perfusion ratios were highest in animals receiving UCB CD133(+) cells, while animals receiving BM CD133(+) cells and BM MNC demonstrated perfusion ratios statistically higher than in animals treated with cytokine media alone. Animals receiving CD133(+) cells showed a statistically higher capillary density, reduced severe digit necrosis and increased engraftment in the BM than animals treated with unselected BM MNC. In vitro studies showed equivalent migration to stromal-derived factor-1 (SDF-1), increased production of tumor necrosis factor alpha (TNF-alpha) and increased branch points with the co-incubation of CD133(+) cells with human umbilical vein endothelial cells (HUVEC) in the Matrigel angiogenesis assay.
Taken together, UCB CD133(+) cells exhibit robust vasculogenic functionality compared with BM MNC in response to ischemia.
Cytotherapy 11/2009; 12(1):67-78. · 3.63 Impact Factor
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ABSTRACT: The stem cell therapy for treating ischemic diseases is promising; however, the limited availability and compromised quality of progenitor cells in aged and diseased patients limit its therapeutic use. Here we report a nanofiber-based ex vivo stem cell expansion technology and proangiogenic growth factors overexpression of human umbilical cord blood (UCB)-derived progenitor cells to enhance angiogenic potential of therapeutic stem cells. The progenitor cells were expanded approximately 225-fold on nanofiber-based serum-free ex vivo expansion culture technique without inducing differentiation. The expanded cells express high levels of stem cell homing receptor, CXCR4, and adhesion molecule, LFA-1. The nanofiber-expanded stem cells uptake AcLDL effectively, and migrate efficiently in an in vitro transmigration assay. These expanded cells can also differentiate into endothelial and smooth muscle cells in vitro. In a NOD/SCID mouse hind limb vascular injury model, nanofiber-expanded cells were more effective in blood flow restoration and this effect was further augmented by VEGF(164) and PDGF-BB, growth factor overexpression. The data indicate that nanofiber-based ex vivo expansion technology can provide an essential number of therapeutic stem cells. Additionally, proangiogenic growth factors overexpression in progenitor cells can potentially improve autologous or allogeneic stem cell therapy for ischemic diseases.
Cell Transplantation 02/2009; 18(3):305-18. · 5.13 Impact Factor
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Hiranmoy Das,
Jon C George,
Matthew Joseph,
Manjusri Das,
Nasreen Abdulhameed,
Anna Blitz,
Mahmood Khan,
Ramasamy Sakthivel,
Hai-Quan Mao,
Brian D Hoit,
Periannan Kuppusamy, Vincent J Pompili
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ABSTRACT: Therapeutic potential was evaluated in a rat model of myocardial infarction using nanofiber-expanded human cord blood derived hematopoietic stem cells (CD133+/CD34+) genetically modified with VEGF plus PDGF genes (VIP).
Myocardial function was monitored every two weeks up to six weeks after therapy. Echocardiography revealed time dependent improvement of left ventricular function evaluated by M-mode, fractional shortening, anterior wall tissue velocity, wall motion score index, strain and strain rate in animals treated with VEGF plus PDGF overexpressed stem cells (VIP) compared to nanofiber expanded cells (Exp), freshly isolated cells (FCB) or media control (Media). Improvement observed was as follows: VIP>Exp> FCB>media. Similar trend was noticed in the exercise capacity of rats on a treadmill. These findings correlated with significantly increased neovascularization in ischemic tissue and markedly reduced infarct area in animals in the VIP group. Stem cells in addition to their usual homing sites such as lung, spleen, bone marrow and liver, also migrated to sites of myocardial ischemia. The improvement of cardiac function correlated with expression of heart tissue connexin 43, a gap junctional protein, and heart tissue angiogenesis related protein molecules like VEGF, pNOS3, NOS2 and GSK3. There was no evidence of upregulation in the molecules of oncogenic potential in genetically modified or other stem cell therapy groups.
Regenerative therapy using nanofiber-expanded hematopoietic stem cells with overexpression of VEGF and PDGF has a favorable impact on the improvement of rat myocardial function accompanied by upregulation of tissue connexin 43 and pro-angiogenic molecules after infarction.
PLoS ONE 01/2009; 4(10):e7325. · 4.09 Impact Factor
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ABSTRACT: Clinical trials using intracoronary (IC) delivery of cells have addressed efficacy but the optimal delivery technique is unknown. Our study aimed to determine whether transvenous intramyocardial (TVIM) approach was advantageous for cellular retention in AMI.
Domestic pigs (n = 4) underwent catheterization with coronary angiography and ventriculography prior to infarction and pre- and post-cells. Pigs underwent 90-minute balloon occlusion of the left anterior descending artery (LAD). After one week they were prepared for IC (n = 2) or TVIM (n = 2) delivery of bone marrow mononuclear cells (MNC) labeled with GFP. IC infusion used an over-the-wire catheter to engage the LAD and balloon inflation to prevent retrograde flow. Venography via the coronary sinus was used for TVIM delivery. The anterior interventricular vein was engaged with a guidewire allowing use of the TransAccess catheter that is outfitted with an ultrasound tip for visualization. Animals were sacrificed one hour after delivery and tissue was analyzed.
Procedures were performed without complication and monitoring was uneventful. 1 x 10(8) MNC were isolated from each bone marrow (BM) preparation and 1 x 10(7) MNC delivered. Ventriculography at one week revealed wall motion abnormalities consistent with an anterior AMI. TVIM and IC delivery revealed mean 452 cells per section and 235 cells per section on average, respectively, in the infarct zone (P = 0.01).
We have demonstrated that TVIM approach for cell delivery is feasible and safe. Moreover, this approach may provide an advantage over IC infusion in retention of the cellular product; however, larger studies will be necessary.
Journal of Interventional Cardiology 11/2008; 21(5):424-31. · 1.18 Impact Factor
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ABSTRACT: The treatment of cardiovascular disease has benefited from advances in pharmacologic and intravascular intervention reducing the morbidity and mortality associated with this disease. To address the need in managing clinically complex vascular disease with limited therapeutic options studies have focused on cellular therapy as a means to augment compensatory mechanisms and to potentially prevent escalation and advancement of disease. Umbilical cord blood (UCB) is a rich source of hematopoietic stem cells (HSC) and thus may be a potential source of cells for this type of therapy. UCB can be collected at no risk to the donor, is immediately available, has a wider availability of HLA phenotypes with a possible lower immune reactivity and does not provoke ethically charged debates. Moreover, stem cells isolated from patients with chronic disease have impairment of their reparative abilities thus limiting their therapeutic impact. The potential of UCB HSC in augmenting this process has been studied extensively both in vitro and in vivo and has shown a benefit in acute and chronic vascular ischemia. Although studies suggest efficacy with no obvious safety concerns the mechanism for this therapeutic effect is unknown.
Journal of Molecular and Cellular Cardiology 06/2007; 42(5):912-20. · 5.17 Impact Factor
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ABSTRACT: Intracoronary mononuclear cell therapy may produce angiogenesis in chronic myocardial ischemia. Potential complications include periprocedural infarction secondary to: reduced coronary blood flow; hyperviscosity from the cell preparation; or microvascular dysfunction. To date, no studies to evaluate these potential complications have been reported. The objective of this report was to study the safety and feasibility of intracoronary injections of autologous bone marrow mononuclear cells in a porcine chronic myocardial ischemia model.
Domestic pigs (n = 5) underwent ameroid cuff placement of the left circumflex artery. Bone marrow-derived mononuclear cells [15 x 10(6) cells] labeled with CM dioctadecyl tetramethylindocarbocyanine were given by intracoronary injection. Animals were sacrificed, and hearts and vital organs were inspected grossly and by histopathology, and bone marrow underwent immunofluorescence microscopy.
Troponin I levels, gross inspection and histopathology did not reveal evidence of myocardial infarction. Labeled cells were observed in perivascular structures in myocardium at the injection site in all animals and in the spleen from one animal. Bone marrow aspirates indicated labeled cells.
Intracoronary injection of autologous mononuclear cells in a porcine chronic myocardial ischemia model appears safe. Intracoronary injection resulted in cell localization in the perivascular areas of myocardium supplied by the injected vessel. Cell localization was observed only in the spleen in just one animal. Labeled cells were identified in bone marrow aspirates from three animals following injection, suggesting a role for bone marrow engraftment and repopulation as a possible mechanism for progenitor cell localization in myocardium.
The Journal of invasive cardiology 06/2006; 18(5):212-8. · 1.84 Impact Factor
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Marcie R Finney,
Nicholas J Greco,
Stephen E Haynesworth,
Joseph M Martin,
David P Hedrick,
Jimmy Z Swan,
Daniel G Winter,
Suzanne Kadereit,
Matthew E Joseph,
Pingfu Fu, Vincent J Pompili,
Mary J Laughlin
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ABSTRACT: Endothelial precursor cells (EPCs) cultured from adult bone marrow (BM) have been shown to mediate neovasculogenesis in murine models of vascular injury. We sought to directly compare umbilical cord blood (UCB)- and BM-derived EPC surface phenotypes and in vivo functional capacity. UCB and BM EPCs derived from mononuclear cells (MNC) were phenotyped by surface staining for expression of stromal (Stro-1, CXCR4, CD105, and CD73), endothelial (CD31, CD146, and vascular endothelial [VE]-cadherin), stem cell (CD34 and CD133), and monocyte (CD14) surface markers and analyzed by flow cytometry. The nonobese diabetic/severe combined immunodeficiency murine model of hind-limb ischemia was used to analyze the potential of MNCs and culture-derived EPCs from UCB and BM to mediate neovasculogenesis. Histologic evaluation of the in vivo studies included capillary density as a measure of neovascularization. Surface CXCR4 expression was notably higher on UCB-derived EPCs (64.29%+/-7.41%) compared with BM (19.69%+/-5.49%; P=.021). Although the 2 sources of EPCs were comparable in expression of endothelial and monocyte markers, BM-derived EPCs contained higher proportions of cells expressing stromal cell markers (CD105 and CD73). Injection of UCB- or BM-derived EPCs resulted in significantly improved perfusion as measured by laser Doppler imaging at days 7 and 14 after femoral artery ligation in nonobese diabetic/severe combined immunodeficiency mice compared with controls (P<.05). Injection of uncultured MNCs from BM or UCB showed no significant difference from control mice (P=.119; P=.177). Tissue samples harvested from the lower calf muscle at day 28 demonstrated increased capillary densities in mice receiving BM- or UCB-derived EPCs. In conclusion, we found that UCB and BM-derived EPCs differ in CXCR4 expression and stromal surface markers but mediate equivalent neovasculogenesis in vivo as measured by Doppler flow and histologic analyses.
Biology of Blood and Marrow Transplantation 05/2006; 12(5):585-93. · 3.87 Impact Factor
