Biomedical subjects
Publications and source records attributed to Nancy Boudreau.
One goal of biomedical research is to reliably construct surrogate tissues for replacement therapy and to promote tissue regeneration. In this issue of Cell, Chun et al. (2006) provide insight into the molecular basis of tissue-specific differentiation. The authors show that remodeling of the extracellular matrix by the matrix metalloproteinase MT1-MMP contributes to the three-dimensional development of white adipose tissue in mice.
Hemangiomas are the most common benign tumor of childhood. Clinical management is limited primarily to observation. Non-surgical treatment modalities have had mixed results and with morbid side effects. Improved understanding of angiogenesis over the last two decades is helping to delineate differences between various vascular tumors. This molecular understanding will be central in helping to properly diagnose and potentially treat these childhood tumors. While a number of downstream effector cytokines have been shown to have altered expression in hemangiomas, a cause for the primary dysregulation within hemangiomas has not yet been clarified. Upstream modulators of angiogenesis are now being defined. Homeobox (Hox) genes are master transcription factors, which have a centrol role during organogenesis, and more recently have been documented to be involved in postnatal tissue remodeling and tumor angiogenesis. We document increased expression of Hox D3 in proliferating hemangiomas and propose a potential role for Hox A3, B3, A5 and D10.
Wound repair requires both the recruitment and coordination of numerous cell types including inflammatory cells, fibroblasts, endothelial and epithelial cells. Each cell type has a distinct set of cell behavior such as formation of granulation tissue and basement membrane, migration, proliferation and redifferentiation. These processes are dependent on cell-cell and cell-ECM signaling, intracellular signal transduction cascades, and ultimately, changes in gene transcription. We have investigated the role of the transcription factor HOXA3 in wound repair and angiogenesis. Here we show that HOXA3 increases endothelial cell migration, induces angiogenesis in vivo, and leads to increased expression of the matrix metalloproteinase-14 (MMP-14) and urokinase-type plasminogen activator receptor (uPAR) genes in endothelial cells in culture and in vivo in response to injury. We find that HOXA3 gene expression is upregulated during wound healing in angiogenic endothelial cells and keratinocytes, and that HOXA3 is not induced in genetically diabetic mice that have impaired angiogenesis and wound repair. We demonstrate that gene transfer of HOXA3 into diabetic mouse wounds leads to dramatic improvements in both angiogenesis and wound closure. In addition, we show that HOXA3 promotes migration of endothelial cells and keratinocytes in a uPAR-dependent manner. Together these findings illustrate how the morphoregulatory protein, HOXA3 can facilitate tissue remodeling via coordinated changes in both epithelial and endothelial cell gene expression and behavior in adult tissues during wound repair.
Brain hemorrhage is a severe complication of both neoplastic and nonneoplastic brain disease. Mice deficient in the alpha(v)beta8 integrin display defective brain vessel formation resulting in hemorrhage and perinatal death, but the mechanism of brain hemorrhage is unknown. Because the alpha(v)beta8 integrin is expressed by astrocytes and not expressed by endothelium, paracrine interactions between astrocytes and endothelial cells could contribute to the maintenance of brain vessel integrity. We have investigated the mechanisms underlying astrocytic-endothelial paracrine signaling and have found that integrin-mediated activation of transforming growth factor (TGF)-beta by astrocytes influences endothelial cell function. Thus, we identified the integrin alpha(v)beta8 in human perivascular glial cell processes surrounding developing blood vessels. Human astrocytic alpha(v)beta8 was a major cell surface receptor for latent TGF-beta, and alpha(v)beta8-dependent activation of TGF-beta was the major mechanism of TGF-beta activation in primary cultures of astrocytes or freshly dissociated fetal brain cells. This activation of TGF-beta was sufficient to inhibit endothelial migration in fibrin gels and to alter expression of genes affecting proteolytic and angiogenic pathways. Taken together, our data suggest that astrocytic alpha(v)beta8 acts as a central regulator of brain vessel homeostasis through regulation of TGF-beta activation and expression of TGF-beta-responsive genes that promote vessel differentiation and stabilization, most notably plasminogen activator inhibitor-1 and thrombospondin-1.
BACKGROUND: Homeobox (Hox) genes are transcriptional regulators which modulate embryonic morphogenesis and pathological tissue remodeling in adults via regulation of genes associated with cell-cell or cell extracellular matrix (ECM) interactions. We previously showed that while Hox 3 genes promote angiogenesis, Hox D10 inhibits this process. METHODS AND RESULTS: Here we show that another Hox family gene, Hox A5, also blocks angiogenesis but accomplishes this by targeting different downstream genes than Hox D10. Sustained expression of Hox A5 leads to down regulation of many pro-angiogenic genes including VEGFR2, ephrin A1, Hif1alpha and COX-2. In addition, Hox A5 also upregulates expression of anti-angiogenic genes including Thrombospondin-2. Furthermore, we show that while Hox A5 mRNA is expressed in quiescent endothelial cells (EC), its expression is diminished or absent in active angiogenic EC found in association with breast tumors or in proliferating infantile hemangiomas. CONCLUSIONS: Together our results suggest that restoring Hox A5 expression may provide a novel means to limit breast tumor growth or expansion of hemangiomas.
Sulf-2 is an endosulfatase with activity against glucosamine-6-sulfate modifications within subregions of intact heparin. The enzyme has the potential to modify the sulfation status of extracellular heparan sulfate proteoglycan (HSPG) glycosaminoglycan chains and thereby to regulate interactions with HSPG-binding proteins. In the present investigation, data mining from published studies was employed to establish Sulf-2 mRNA upregulation in human breast cancer. We further found that cultured breast carcinoma cells expressed Sulf-2 mRNA and released enzymatically active proteins into conditioned medium. In two mouse models of mammary carcinoma, Sulf-2 mRNA was upregulated in comparison to its expression in normal mammary gland. Although mRNA was present in normal tissues, Sulf-2 protein was undetectable; it was, however, detected in some premalignant lesions and in tumors. The protein was localized to the epithelial cells of the tumors. In support of the possible mechanistic relevance of Sulf-2 upregulation in tumors, purified recombinant Sulf-2 promoted angiogenesis in the chick chorioallantoic membrane assay.
INTRODUCTION: Medical diagnostic tests are evaluated based on measures of sensitivity (Sn), specificity (Sp), and likelihood ratios (LR). These procedures are limited in the event of a biased gold standard or missing data. Interpretations of these measures are frequently inappropriate. PURPOSE: The Rasch measurement model (RMM) was examined as a method to provide evidence of diagnostic test utility in order to overcome the limitations of Sn, Sp, and LR. METHODS: Patients suspected of a knee ligament tear (n = 825) were studied, by evaluating four diagnostic tests. The RMM probability estimates for each test were compared to estimates of Sn, Sp, and LR. RESULTS: The RMM provided probability estimates for the diagnosis that were comparable to likelihood ratios. These probability estimates correlated with the estimates of Sn, Sp, and LR. The RMM estimates were not affected by missing data. DISCUSSION: The RMM may provide an alternative means to study the utility of medical diagnostic tests to estimate the probability of disease presence/absence.
Angiogenesis is regulated by concerted actions of angiogenic and angiostatic factors. Homeobox D3 gene (HOXD3) is a potent proangiogenic transcription factor that promotes angiogenesis by modulating the expression of matrix-degrading proteinases, integrins, and extracellular matrix components. Application of HOXD3 can promote angiogenesis in the skin, but its role in other vascular beds has not been examined. The authors examined HOXD3 expression in human brain vessels by in situ hybridization. Although little or no HOXD3 mRNA was detected in normal brain vessels, increased levels of HOXD3 and its target gene, alpha V beta 3, were found in angiogenic vessels in human brain arteriovenous malformations. The authors further investigated whether HOXD3 plays a role in cerebral angiogenesis in a murine model. Expression of HOXD3 in mouse brain was achieved through retroviral vector-mediated HOXD3 gene transfer. HOXD3 expression lead to a significant induction of cerebral angiogenesis as shown by quantitative microvessel counting (HOXD3: 241 +/- 19 vessels/mm2 vs. saline: 150 +/- 14 vessels/mm2, P < 0.05). The data also showed that focal cerebral blood flow was increased in the angiogenic region with less vascular leakage. Moreover, expression of HOXD3 led to an increase in the expression of a direct downstream target gene alpha V beta 3 integrin. The data suggest that HOXD3 may play an important role in regulating cerebral angiogenesis, and that gene transfer of HOXD3 may provide a novel and potent means to stimulate angiogenesis.
Growth and progression of breast cancers are accompanied by increased neovascularization (angiogenesis). A variety of factors, including hypoxia and genetic changes in the tumor cells, contribute to increased production of angiogenic factors. Furthermore, cells within the activated tumor stroma also contribute to the increase in production of vascular endothelial growth factor and other angiogenic factors, including basic fibroblast growth factor and platelet-derived growth factor. The contribution of the microenvironment to tumor-induced angiogenesis is underscored by findings that breast tumors implanted into different tissue sites show marked differences in the extent and nature of the angiogenic response. These findings have important implications for designing anti-angiogenic therapies.
Poorly healing diabetic wounds are characterized by diminished collagen production and impaired angiogenesis. HoxD3, a homeobox transcription factor that promotes angiogenesis and collagen synthesis, is up-regulated during normal wound repair whereas its expression is diminished in poorly healing wounds of the genetically diabetic (db/db) mouse. To determine whether restoring expression of HoxD3 would accelerate diabetic wound healing, we devised a novel method of gene transfer, which incorporates HoxD3 plasmid DNA into a methylcellulose film that is placed on wounds created on db/db mice. The HoxD3 transgene was expressed in endothelial cells, fibroblasts, and keratinocytes of the wounds for up to 10 days. More importantly, a single application of HoxD3 to db/db mice resulted in a statistically significant acceleration of wound closure compared to control-treated wounds. Furthermore, we also observed that the HoxD3-mediated improvement in diabetic wound repair was accompanied by increases in mRNA expression of the HoxD3 target genes, Col1A1 and beta 3-integrin leading to enhanced angiogenesis and collagen deposition in the wounds. Although HoxD3-treated wounds also show improved re-epithelialization as compared to control db/db wounds, this effect was not due to direct stimulation of keratinocyte migration by HoxD3. Finally, we show that despite the dramatic increase in collagen synthesis and deposition in HoxD3-treated wounds, these wounds showed normal remodeling and we found no evidence of abnormal wound healing. These results indicate that HoxD3 may provide a means to directly improve collagen deposition, angiogenesis and closure in poorly healing diabetic wounds.
The ECM protein Del-1 is one of several novel ECM proteins that accumulate around angiogenic blood vessels in embryonic and tumor tissue and promote angiogenesis in the absence of exogenous growth factors. Del-1 expressed in mouse or rabbit ischemic hind-limb muscle by gene transfer rapidly promotes new blood vessel formation and restores muscle function. This angiogenic ECM protein initiates angiogenesis by binding to integrin alphavbeta5 on resting endothelium, thereby resulting in expression of the transcription factor Hox D3 and integrin alphavbeta3. Hox D3 converts resting endothelium to angiogenic endothelium by inducing expression of proangiogenic molecules such as integrin alphavbeta3. These findings provide evidence for an angiogenic switch that can be initiated in the absence of exogenous growth factors and indicate that the angiogenic matrix protein Del-1 may be a useful tool for the therapy of ischemic disease.
Homeobox (Hox) genes are master regulatory genes that direct organogenesis and maintain differentiated tissue function. As HoxD3 and HoxB3 promote angiogenesis, we investigated whether endothelial cells use other Hox genes to maintain a mature quiescent phenotype. HoxD10 expression was higher in quiescent as compared to tumor-associated angiogenic endothelium. Microarray analysis of HoxD10-overexpressing endothelial cells revealed a pattern of gene expression consistent with a nonangiogenic phenotype. Moreover, sustained expression of HoxD10 impaired endothelial cell migration and blocked angiogenesis induced by basic fibroblast growth factor and vascular endothelial growth factor in the chick chorioallantoic membrane in vivo. HoxD10-overexpressing human endothelial cells also failed to form new vessels when implanted into immunocompromised mice. These results indicate a role for HoxD10 in maintaining a nonangiogenic state in the endothelium.
To study a potential feedback system in the angiopoietin (Ang)-Tie2 system, the authors examined effects of Ang1 and Ang2 on Tie2 expression on human umbilical vein endothelial cells (HUVECs) with or without stimulation by a potent inflammatory cytokine, tumor necrosis factor-alpha (TNF-alpha). Ang1, but not Ang2, down-regulated Tie2 expression on HUVECs without TNF-alpha stimulation. Both Ang1 and Ang2 attenuated TNF-alpha-induced Tie2 up-regulation. Regulation of Tie2 expression by Ang1 or Ang2 was not dependent on phosphatidylinositol 3-kinase. The Ang-Tie2 system appears to have an autoregulatory feedback system that may be regulating the overall activity of the Tie2 system in both physiological and pathological conditions.