Angiogenesis, or the formation of new blood vessels from the existing ones, is an essential biological process for maintaining numerous physiological functions ranging from cell growth, proliferation, repair of damaged cells to wound-healing in vivo [1–3]. Throughout the life of an individual and during embryonic development, various pro-angiogenic and anti-angiogenic factors (i.e. promoters and inhibitors of angiogenesis respectively) produced by various cell types maintain a balance between neovascularization and angiogenesis programs in a cyclic manner [4, 5]. Exactly how abnormal angiogenic signals are generated in vivo is not well-understood, but an imbalance in the production of one or more critical factors can alter the protein-protein interaction pathways and induce angiogenic anomalies including inflammation, vascular dementia, hemangioma, dysfunctional uterine bleeding, ovarian hyperstimulation and choroidal/intraocular disorders to name a few [1, 6]. Angiogenesis is also critical for cancer metastasis, diabetic blindness, age-related macular degeneration, rheumatoid arthritis, psoriasis, and for the development of new blood vessels that supply oxygen and nutrients to the body when aortas are clogged (thrombosis) [2, 6].
In both the neoplastic and non-neoplastic diseases, endothelial cells have been shown to express various isoforms of the vascular endothelial growth factors (VEGFs) which bind to their cognate VEGF receptors (VEGFRs), activate their associated protein tyrosine kinases (PTKs) and stimulate endothelial cell growth through angiogenic pathways [3, 6, 7]. However, endothelial cells can be activated by various cytokines, phosphorylated proteins and other factors that are essential not only for cell growth but also for maintaining an activated state of the stimulated endothelial cells [2, 8]. In the absence of specific cytokines and diffusible signaling proteins, VEGF by itself is not sufficient to trigger expression of numerous enzymes and proteins required for the development of a network of blood vessels from the existing vasculature [8, 9].
Angiogenic Factors are also produced by Pathogenic Viruses
Etiologic factors involved in different types of vasculopathies in humans have not been fully explored. However, in the absence of any tumor growth many DNA or RNA viruses have been shown to cause vascular lesions in vivo or produce proangiogenic factors in vitro. For example, the human herpes simplex virus type 1 (HSV-1)-infected ocular cells produce IL-6, which stimulates uninfected, avascular corneal cells to secrete VEGF and provoke neovascularization in the eye . Infection with the Epstein-Barr virus (EBV) enhances production of many cytokines and causes angiogenic cutaneous tumors . The dengue virus, causes hemorrhagic fever and vascular lesions in humans, produces interleukin-4 (IL-4), IL-8, IL-6, IL-10, GM- colony stimulating factor (CSF), interferon-gamma (INF-gamma) and tumor necrosis factor alpha (TNF-alpha) . The human parapoxvirus causes extensive skin vasculopathies and the pseudocowpox viral genome induces viral gene-encoded VEGF homologues (i.e. VEGF-like factors) [13, 14]. Likewise, the common human rhinovirus infection produces factors that promote angiogenesis in bronchial epithelial cells .
One of the best-studied models of angiogenesis is Kaposi's sarcoma (KS), a highly vascular tumor that is rare in the general population but occurs frequently in human immunodeficiency virus (HIV)-infected individuals [16–18]. However, KS is etiologically associated with the human herpesvirus-type-8 (HHV-8) infection since HHV8-genome itself encodes a viral G-protein-coupled receptor (vGPCR), which activates both oncogenic and angiogenic pathways in the presence or absence of HIV-coinfection [17, 19, 20].
Many HIV-infected patients, who may or may not be infected with HHV8, develop intraepithelial neoplasia, hemangiomas, lymphomas, angiosarcomas, myelodysplastic angiogenic syndrome and other angiopathies [21–23]. The HIV-encoded transcriptional transactivator (Tat) protein has been implicated in angiogenesis because it binds VEGFR and stimulates endothelial cell growth . However, its binding-affinity is not as strong as that of the natural cellular VEGFs and the avidity of Tat interaction with VEGFR is dependent on specific cytokines produced locally by endothelial cells, cancer cells or other virus-infected and uninfected cell types in vivo [10, 13, 24, 25]. Further, the activated state of endothelial cells must be maintained continuously during the numerous biological processes that lead to angiogenesis. These data suggest that while Tat synergizes the effects of many viral and cellular factors during the complex biological processes of angiogenesis, Tat alone or individual cytokines by themselves do not induce angiogenesis in mice.
The molecular mechanisms involved in HIV-induced vasculopathies in humans are difficult, if not impossible to study because most patients are co-infected with different pathogenic viruses such as HSV-1, HSV11, EBV, hepatitis B virus (HBV), hepatitis C virus (HCV), human papilloma virus (HPV) and different bacterial and fungal microorganisms. Consequently, cellular changes induced by HIV alone in vivo can not be distinguished from those produced by other viruses or pathogenic organisms co-inhabiting the same individual, unless separate protein profiles of each class of different infectious agents are established first. We therefore tested a hypothesis that chronic HIV-replication in non-endothelial cells induces novel cellular proteins that provoke specific protein-protein interactions along the angiogenic pathways. Although most in vitro studies have utilized endothelial cells derived from early KS lesions or human veins (by necessity), in this study we preferred to use T-cells because some differentiated endothelial cells may already produce proangiogenic cytokines in response to changes in the cellular milieu or alternatively, factors that are essential for endothelial cell activation may be experimentally induced [26, 27]. Herein, we report that HIV- infected human T-cells produce numerous kinases, adhesion molecules and other angiogenic factors (not encoded by HIV-genome) that are capable of initiating and promoting novel VEGF-independent pathways. These mechanisms are similar to those observed during embryonic development, neovascularization and angiogenesis.