Characterization of Lymphocyte-Dependent Angiogenesis Using a SCID Mouse: Human Skin Model of Psoriasis

Characterization of Lymphocyte-Dependent Angiogenesis Using a SCID Mouse: Human Skin Model of Psoriasis Brian J. Nickoloff

Department of Pathology, Skin Cancer Research Laboratories, Cardinal Bernardin Cancer Center, Loyola University Medical Center, Maywood, Illinois, U.S.A.

skin engrafted onto severe combined immunode®cient mice followed by injection of activated immunocytes. This new experimental model represents a reproducible and pharmacologically validated method to trigger neovascularization and bona ®de psoriatic plaque formation. In addition, the potential contribution of epidermal keratinocytes and dermal macrophages to the angiogenic tissue reaction is presented, and a series of questions are then posed that can be answered using the severe combined immunode®cient mouse model of psoriasis. Finally, a model is proposed integrating all available data into a coherent multistep reaction schema that includes active participation by multiple cell types including natural killer T cells, keratinocytes, macrophages, and microvascular endothelial cells. Key words: endothelial cells/immunocytes/macrophages/T cells. Journal of Investigative Dermatology Symposium Proceedings 5:67±73, 2000

From a clinical perspective, angiogenesis is an important component of acute and chronic psoriatic skin lesions as they are erythematous and display a tendency to bleed after super®cial removal of scale. By routine histology, numerous microscopic vascular abnormalities are also present. The structural expansion of capillaries and distinctive activated phenotype of lesional endothelial cells are believed not only to be clinical and pathologic hallmarks of the disease, but to play a central role in the pathogenesis of psoriatic plaques. Despite over 20 years of research by leading angiogenesis experts and numerous studies, many details regarding the cellular and molecular basis for angiogenesis in psoriasis remain unknown. In this review, 10 different sections are presented to update recent progress in this active ®eld of investigative skin biology. Highlights of this review include the phenotypic characterization of endothelial cells in acute and chronic psoriatic plaques, and a review of a novel animal model of psoriasis using human

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tissue reaction (Fig 1B). In dozens of injected grafts, psoriatic plaques created in this model system are always accompanied by a prominent angiogenic tissue reaction. There are many changes in the blood vessels within a chronic, well-established psoriatic plaque including: dilation of capillary beds, increased capillary permeability, increased endothelial cell proliferation, and tortuoisity of capillary loops with increase in blood ¯ow through the skin (Braverman and Yen, 1977; Ryan, 1980; Braverman and Sibley, 1982; Klemp and Staberg, 1983; Heng et al, 1991). The study of the vascular changes in psoriatic plaques has been of interest to many leading scientists interested in angiogenesis for several decades (Folkman, 1972, 1995). By using the SCID mouse model, it is possible to determine the earliest molecular and cellular changes in the epidermis and dermis following injection of activated immunocytes that lead to the characteristic angiogenic tissue response observed in psoriasis. Previous investigators have observed that peripheral bloodderived mononuclear cells from psoriatic patients, or serum from psoriatic patients, could induce angiogenesis (Majewski et al, 1985, 1987); however, the exact cell-type (i.e., T cell subset), or the molecules responsible for this angiogenic response, were not further characterized. More recently, attempts have been made to de®ne the cytokine network in psoriasis, and to identify angiogenic as well as angiostatic cytokines that may contribute to the vascular changes present in psoriatic plaques (Nickoloff, 1991). In this review, a summary of the literature concerning the molecular and cellular

soriasis is a chronic in¯ammatory disease characterized by prominent epidermal proliferation (Wrone-Smith and Nickoloff, 1996). Using a novel severe combined immunode®cient (SCID) mouse:human skin chimeric animal model, it has been established that activated immunocytes are responsible for triggering the keratinocyte hyperplasia producing a thick epidermis with altered differentiation and scale production (Nickoloff et al, 1995; Boehncke et al, 1996; Gilhar et al, 1997; Sugai et al, 1998; Yamamoto et al, 1998). In addition to activated T cells and natural killer T cells being involved in provoking rapid and substantial keratinocyte hyperplasia, the engrafted skin samples also consistently display rather remarkable and rapid new blood vessel formation (i.e., angiogenesis) following injection of activated immunocytes (Boehncke et al, 1996; Nickoloff et al, 1999). When engrafted prepsoriatic skin is injected with saline, the dermal vascularization in the human dermis contains only relatively inconspicuous blood vessels (Fig 1A), but within 2 weeks of injection of activated immunocytes, the human dermis contains numerous blood vessels representing an angiogenic Manuscript received January 28, 2000; revised April 18, 2000; accepted for publication May 25, 2000. Reprint requests to: Dr. Brian J. Nickoloff, Director, Skin Cancer Research Program, Cardinal Bernardin Cancer Center, Building #112, Room 301, 2160 South First Avenue, Maywood, IL 60153. Email: [email protected] 1087-0024/00/$15.00

´ Copyright # 2000 by The Society for Investigative Dermatology, Inc. 67

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Figure 1. Angiogenic tissue reaction in engrafted prepsoriatic human skin transplanted onto SCID mice. When orthotopically transplanted skin is injected with saline, no angiogenesis is present (A), but 2 weeks after injection of activated immunocytes the human dermis contains numerous small and large blood vessels re¯ecting a local angiogenic tissue response that accompanies the other clinical and microscopic changes characteristic of a bona ®de plaque of psoriasis (B).

Figure 2. Differential expression of avb3 versus avb5 integrins in NN, PN, and chronic psoriatic plaques (PP skin), as well as acute psoriatic lesions created using the SCID mouse model. Immunoperoxidase staining; Vectastain (anti-avb3 integrin MoAb; MoAb1976, and anti-aVb5 integrin MoAb; MoAb1961; anti-ELAM-1; CD62E). 3-amino-4-ethylcarbazole as chromagen producing positive red reaction product. (A) avb3 expression is absent in NN skin, but is focally present in PN skin and diffusely positive in endothelial cells in chronic PP skin. An acute psoriatic lesion created by injecting activated autologous immunocytes into engrafted PN skin reveals endothelial cell positivity for avb3 integrin (panel labelled SCID). (B) avb5 expression is focally present on upper dermal dendritic cells, and on basal keratinocytes and dermal dendritic cells in PN skin. In a chronic psoriatic plaque, while perivascular and interstitial dendritic cells are avb5 positive, the endothelial cells are negative. An acute psoriatic lesion created on a SCID mouse (far right panel) has strong and diffuse dendritic cell avb5 expression, but no endothelial cell positivity. Inset reveals ELAM-1 expression on microvascular endothelial cells.

basis for angiogenesis in psoriasis will be followed by a brief discussion of currently unresolved questions, and a model for the pathogenesis of psoriasis integrating all available data will conclude the presentation. CHARACTERIZATION OF PHENOTYPE OF ACTIVATED ENDOTHELIAL CELLS IN CHRONIC PSORIATIC PLAQUES From a clinical and morphologic perspective, the prominent vasculature in the super®cial plexus of a chronic psoriatic plaque point to endothelial cell proliferation and activation (Wolfe, 1989).

To characterize the phenotype of the activated endothelial cell, investigators have used immunohistochemical staining and compared various antigens expressed by endothelial cells of normal skin obtained from healthy individuals (NN skin), with clinically symptomless skin removed from patients with psoriasis elsewhere (PN skin), as well as chronic psoriatic plaques (PP skin). Because the angiogenic pathway by which early endothelial cells sprout and extend to form tubes and elongate capillary beds involves interactions with extracellular matrix proteins, various integrins have been examined (Ingber, 1992). Integrins are cell surface molecules that play key roles in the angiogenic process because they mediate cell±matrix interaction and trigger intracellular signaling

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Figure 3. Psoriatic plaque contains a rich admixture of macrophages and dendritic cells. The macrophage population includes a pan-macrophage marker CD14, macrophages with classical activation markers (i.e., CD16, CD32) as well as alternatively activated macrophages (CD163, mannose receptor). In addition, there are epidermal and dermal dendritic cells expressing CD1a, CD80, and CD83, indicating various degrees of maturation of these professional antigen-presenting cells.

producing endothelial cell activation and proliferation (Hynes, 1992). In NN skin and PN skin there was low to absent expression of the avb3 integrin (Brooks et al, 1994), but in psoriatic plaques there was strong and diffuse endothelial cell expression of avb3 integrin (Creamer et al, 1995). In our own series of six different skin biopsies, we con®rmed these observations with respect to avb3 integrin expression (Fig 2). In contrast to this pattern for avb3 integrin expression, there is actually a decrease in expression of b4 integrin in PP skin compared with PN skin, and no change in b1 integrin expression between these different skin samples (Creamer et al, 1995). Because cytokines can induce angiogenesis, Cheresh et al have de®ned at least two different angiogenic pathways by differential endothelial cell expression of avb3 versus avb5 integrins (Friedlander et al, 1995). This group reported that avb3-dependent angiogenesis was induced by basic ®broblast growth factor (bFGF) or tumor necrosis factor alpha (TNF-a), whereas avb5-dependent angiogenesis was triggered by either vascular endothelial growth factor (VEGF), or transforming growth factor-a (TGF-a), or phorbol ester (Friedlander et al, 1995). To examine the different expressions of avb3 versus avb5, the same six NN, PN, and PP skin biopsies were immunostained. Figure 2 reveals that while there was focal basal keratinocytes and dermal dendritic cell expression of avb5 in NN and PN skin, there was no detectable endothelial cell positivity. Moreover in all PP skin biopsies there was more extensive dermal dendritic cell avb5 expression, but no endothelial cell positivity. The lack of endothelial cell avb5 was rather surprising because the cytokine network in psoriatic plaques includes the presence of VEGF and

TGF-a (inducers of avb5 in endothelial cells). We will return to this topic later in the paper. Before concluding this section, it should also be noted that there are several other phenotypic characteristics that distinguish endothelial cells in PP skin from endothelial cells in NN and PN skin. Other surface antigens prominently expressed by endothelial cells in PP skin include: endothelial cells adhesion molecule-1 (ELAM-1; also called E-selectin) (Petzelbauer et al, 1994), intercellular adhesion molecule-1 (ICAM-1) (Groves et al, 1993), vascular cell adhesion molecule-1 (VCAM-1) (Grif®ths et al, 1989), and the MS-1 antigen (Goerdt et al, 1991). Thus, it is clear that the endothelial cells express a wide variety of markers of activation that distinguish them in the angiogenic tissue reaction characteristic of chronic psoriatic plaques. SWITCHING ON ANGIOGENESIS DURING WOUND HEALING Whereas the previous section focused on chronic psoriatic plaques, in this section the focus will be on describing the dynamic sequential events as recorded during cutaneous wound healing. These types of experiments are relevant to the biology of psoriasis as the ®nal psoriatic plaque may actually represent an abnormal wound-healing response (Lingen and Nickoloff, 1999). There are many different types of wound-healing models in which to study the molecular and cellular basis of the regulation of angiogenesis, but only three models will be discussed; one dealing with porcine skin and the other two using human skin. In the porcine model, the initiation of angiogenesis triggered by full-thickness wounding

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Figure 4. Model for understanding the cellular and molecular basis for the angiogenic tissue reaction in psoriasis highlighting the role of activated immunocytes producing cytokines that trigger neovascularization of the dermis. The upper panels depict the microscopic and clinical appearance of symptomless skin engrafted onto a SCID mouse, and the subsequent psoriatic plaque that is created following injection of activated immunocytes (inset includes prominent neovascularization of dermis underlying newly created psoriatic plaque). The lower panels represent a model in which the intradermal injected activated immunocytes (i.e., natural killer T cells) produce cytokines such as IFN-g that are accompanied by several keratinocyte-derived, macrophage-derived, and dermal dendrocyte-derived pro-angiogenic factors. The net result of this cytokine network is activation and proliferation of aVb3 integrin positive endothelial cells.

revealed the transient functional involvement of the avb3 integrin by endothelial cells, but not avb5 or b1 integrins (Clark et al, 1996). When NN skin was engrafted onto SCID mice, a longitudinal dermo±epidermal excisional wound was created and the subsequent wound healing and angiogenesis process was investigated (Christo®dou-Solonidou et al, 1997). As early as 2 days after wounding, there was upregulation of avb3, avb5, and avb6 integrins by the proliferating endothelial cells, but only inhibitors of avb3 integrin blocked new vessel formation during human wound healing (Christo®dou-Solonidou et al, 1997). To de®ne the primary mediators of angiogenesis in acute wound repair, surgical wound ¯uid was collected from surgical patients undergoing mastectomy or neck dissection from postproductive days 1±7 (Nissen et al, 1998). It was determined that the initial angiogenic stimulus was provided by bFGF-2, which was followed by a more sustained pro-angiogenic stimulus elicited by VEGF (Nissen et al, 1998). The early appearance of bFGF-2 that peaked almost immediately after surgery was presumably released from cells or sequestered by extracellular matrix, and could account for the subsequent upregulation of avb3 integrin expression as noted earlier by the investigators. PHENOTYPE OF DERMAL TISSUE REACTION DURING GENESIS OF PSORIATIC LESIONS Up until recently, it was dif®cult to reproducibly induce and study the onset of psoriasis because no suitable animal model was available; however, with the advent of transplantation technology and availability of SCID mice, an animal model using entirely human skin components was developed and validated (Nickoloff et al, 1995). It is now possible to inject either engrafted PN or NN skin and create psoriatic plaques that include a prominent angiogenic tissue response (Boehncke et al, 1996; Nickoloff et al, 1999). This neovascularization of human dermis takes place within 2±4 weeks following intradermal injection of activated immunocytes (Nickoloff et al, 1999). By using this experimental model system we have begun to characterize the phenotype of the acutely activated and proliferating human dermal microvascular endothelial cells. Figure 2 reveals that acute lesion of psoriasis includes endothelial cells that express avb3, but not avb5 integrins. In addition, note the expression of ELAM-1 by the endothelial cells (Fig 2, inset). Thus, as regards the differential expression of integrins, the acutely generated angiogenic tissue response is similar to chronic psoriatic plaques with respect to the phenotype of the vascular endothelial cells. Now that we can use this experimental approach in which prominent angiogenesis is rapidly and predictably induced by the

injection of activated immunocytes including T cells and natural killer T cells, it will now be possible to more fully explore the molecular mechanisms governing this remarkable change in the clinical and histologic appearance of the skin (Boehncke et al, 1996; Nickoloff and Wrone-Smith, 1999; Nickoloff et al, 1999). POTENTIAL DIRECT MEDIATORS PRODUCED BY ACTIVATED T CELLS THAT CAN MODULATE ANGIOGENESIS Perhaps the simplest view of how the injection of activated immunocyte triggers psoriasis is to consider a direct interaction by which T cells themselves can serve as the source of pro-angiogenic stimuli. This is a reasonable and straightforward possibility as it is well documented that activated T cells can produce pro-angiogenic responses by endothelial cells. Examples of such T cell-derived factors include various cytokines (Lingen and Nickoloff, 1999), as well as scatter factor (hepatocyte growth factor) (Naidu et al, 1994). Scatter factor is a potent angiogenic factor that is present in psoriatic plaques (Grant et al, 1993). Besides pro-angiogenic cytokines, it should also be remembered that T cells in psoriatic plaques also produce cytokines that have direct antiangiogenic effects such as gamma interferon (IFN-g) and interferon inducible protein 10 (IP10) (Reaman and Tosato, 1995). IFN-g has pleiotropic effects (Boehm et al, 1997), as it also is a potent inducer of nitric oxide that can have profound effects on the microvasculature including prominent vasodilation (Xie et al, 1993; Kolb-Bachofen et al, 1994). Thus, it should be clear that there are both positive and negative regulators of microvasculature growth, and that the overall balance amongst these factors determines the ®nal net result with each tissue site of interest (Folkman, 1995). ACTIVATED T CELLS CAN INFLUENCE ENDOTHELIAL CELLS INDIRECTLY VIA THE EPIDERMAL KERATINOCYTE Prior to the discovery of cytokines, epidermal keratinocytes were viewed as rather immunologically inert cells, primarily relegated to a brick and mortar function as regards the barrier function of skin. When T cells were present in the skin, keratinocytes were portrayed as simple passive targets not capable of either initiating or perpetuating in¯ammatory or immune-mediated reactions (Barker et al, 1991). It is now clear, however, that keratinocytes are important coconspirators with T cells and dendritic cells, and can even function as nonprofessional antigen presenting cells (Nickoloff and Turka, 1993). The ability of keratinocytes to produce various cytokines, chemotactic polypeptides, and adhesion molecules has

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led to the recognition that they can in¯uence events in dermis including the activation of endothelial cells (Grif®ths and Nickoloff, 1991; Detmar et al, 1995; Reaman and Tosato, 1995). In this section we will discuss the series of reciprocal interactions by which activated T cells and natural killer T cells move from the dermis into the epidermis and release cytokines that activate keratinocytes, which in turn release additional cytokines that can then activate dermal microvascular endothelial cells in psoriasis. The two most important primary cytokines in psoriatic plaques are IFN-g and TNF-a (Nickoloff, 1991). These cytokines can in¯uence keratinocytes to produce a wide array of other cytokines that have angiogenic activity. Examples of pro-angiogenic regulators produced by cytokine activated keratinocytes include: transforming growth factor-alpha (TGF-a) (Gottlieb et al, 1988; Elder et al, 1989), VEGF (Detmar et al, 1994), IL-8 (Schroder and Christophers, 1986; Nickoloff et al, 1991), scatter factor (Grant DS et al, 1993), and TNF-a (Ettchad et al, 1994). In addition to these pro-angiogenic cytokines, psoriatic keratinocytes underproduce the angioinhibitory molecule thrombospondin (Nickoloff et al, 1994). Given the mitogenic activity of several of these cytokines to also promote further keratinocyte hyperplasia, which can contribute to elongation of psoriatic capillaries (Bacharach-Buchies et al, 1994), and the net pro-angiogenic pro®le of this cytokine network, it is clear that keratinocytes activated by T cells can promote the vascular proliferation that is a hallmark of the psoriatic plaque.

does not express these markers, but does express CD163 and the macrophage mannose receptor (MMR) (Goerdt and Orfanos, 1999). One previous report demonstrated the presence of alternatively activated macrophages in psoriasis, but did not examine the same biopsies for evidence of classical activated macrophages or dendritic cells (Djemadji-Oudjel et al, 1996). To determine if alternatively activated macrophages are the dominant macrophage population in psoriasis, we examined a series of psoriatic plaques with a broad panel of monoclonal antibodies. Because alternatively activated macrophages have been linked to angiogenic responses (Goerdt and Orfanos, 1999), we were intrigued by the possible role for such cells in psoriasis. As shown in Fig 3, a chronic psoriatic plaque contains a rich admixture of dendritic cells and macrophage-appearing cells in the upper dermis immunoreactive for the classical activation markers CD16 and CD32. In addition, there are also numerous positively stained mononuclear cells with alternative activation markers CD163 and MMR. Thus it appears that both types of activated cell phenotypes are present in psoriatic plaques. Once activated, macrophages can induce prominent and rapid angiogenesis by secreting various mediators including TNF-a, TGF-a, IGF-1, PDGF-B, TGF-b, and scatter factor (Leibovich et al, 1987; Kodelja et al, 1997). Further studies are indicated to determine their relative importance and functional contribution to the in¯ammatory process and angiogenic tissue reaction.

ACTIVATED T CELLS AND NATURAL KILLER T CELL INTERACTION WITH MACROPHAGES THAT TRIGGERS ANGIOGENESIS

MODEL LINKING THE GENETIC AND CELLULAR FACTORS RESPONSIBLE FOR TRIGGERING THE ANGIOGENIC TISSUE REACTION IN PSORIASIS

One of the earliest cellular interactions to occur in the skin during the onset of psoriatic lesions involves lymphocytes and macrophages (Schubert and Christophers, 1985). There are also important reactions between T cells and dermal dendritic cells that contribute to the release of cytokines that impact endothelial cells (Nestle et al, 1994). Numerous investigators have documented the presence of various types of monocyte/macrophage and dendritic cells in and around the blood vessels of psoriatic plaques (Paukkonen et al, 1992; Goerdt et al, 1993; Schopf et al, 1993; Sterry and Boehncke, 1993; Oord Van Den and Wolf-Peeters, 1994; Djemadji-Oudjel et al, 1996). Rather than reviewing how the local immune reaction is generated involving antigen presenting cells and T cells, in this section a brief review of the importance of how T cells can impact macrophages, followed by how these activated macrophages can impact endothelial cells will be highlighted. Up until recently, macrophages and dendritic cells were believed to be of distinct origins with nonoverlapping phenotypes and functions (Banchereau and Steinman, 1998); however, it is now clear that monocytes can give rise to dendritic cells and that dendritic cells can undergo phagocytosis (the term dendrophage was introduced to signify this point) (Nickoloff and Nestle, 1995). Currently, it is also possible to delineate at least two different pathways for the activation of macrophages as well as dendritic cells (Goerdt and Orfanos, 1999). These pathways have been identi®ed as either involving classical activation versus alternative activation(s). It has been suggested that acute in¯ammatory processes may be perpetuated into chronic conditions because of these antigen presenting cells (i.e., macrophage and dendritic cells) (Goerdt and Orfanos, 1999). The classical activation pathway involves cytokines such as IL-1, IL-6, IL-12, IFN-g, and TNF-a whereas the alternative activation pathway involves the TH2 type cytokines IL4 and IL-10. The classical macrophage pathways mediates immunologic reactions that produce in¯ammation, whereas the alternative macrophage pathway is envisioned as a counterregulatory process predominantly manifesting itself with downregulation of in¯ammation, angiogenesis, and elimination of tissue debris via phagocytosis as occurs during wound healing. A classical activation-type macrophage has a distinctive immunophenotypic pro®le of cell surface markers including CD16, CD32, and CD64; whereas the alternatively activated macrophage

A line of inquiry that is currently being followed includes the possibility that when one of our natural killer T cell lines is injected into the dermis, because these natural killer T cells can produce both IFN-g (classical activation pathways) as well as IL-4 and IL-13 (alternative activating pathway), neither opposing pathways can become dominantly established within the ®rst 2 weeks (Nickoloff et al 1999). We postulate this potentially unbalanced, and hence dysfunctional interaction, ultimately results in a chronic in¯ammatory reaction with neovascularization that persists for a long period of time. Our current working model includes several sequential cellular and molecular events including the following (Fig 4). First, natural killer T cells in®ltrate the dermis and, upon contact with relevant CD1d bearing target cells such as epidermal keratinocytes, become activated via their natural killer receptors such as CD161 (Bonish et al, 2000). Second, these natural killer T cells begin producing a mixture of cytokines including IFN-g and IL-13 (Nickoloff et al, 2000). Third, the avascular epidermal-based keratinocytes also become activated to release several pro-angiogenic cytokines that in¯uence the underlying vessels in a paracrine fashion. Fourth, the local production of IFN-g and IL-13 activate tissue monocytes/macrophages/dendritic cells and drive both classical as well as alternative activation pathways. Finally, the tissue ``dendrophage'' release a variety of cytokines and angiogenic molecules that provoke a local angiogenic tissue reaction. The proliferating endothelial cells are characterized by a distinctive phenotype including avb3 integrin expression as well as various adhesion molecules including ELAM-1, VCAM-1, and ICAM-1. Thus, the initial in¯ux of pathogenic immunocytes triggers a chain reaction of cellular and molecular events involving both epidermal keratinocytes as well as angiocentric and interstitial dermal macrophages and dendritic cells. It should be noted that the experimental evidence for this new concept in which natural killer T cells can trigger angiogenesis is still preliminary, and will require additional study for comparison with more conventional T cell subsets. REMAINING QUESTIONS THAT CAN BE ADDRESSED USING THE SCID MOUSE ANIMAL MODEL OF PSORIASIS Perhaps the most important currently unsolved issues as regards angiogenesis and psoriasis is whether the neovascularization is required for the creation and/or maintenance of the plaque. Using

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the SCID mouse:human skin chimeric model it is now possible to address this question by preinjecting the engrafted skin with blocking antibodies against the most important integrins such as avb3, followed by intradermal inoculation of activated immunocytes. Alternatively, it is possible to add neutralizing antibody against VEGF to determine the relative importance of angiogenesis to psoriasis mediated by VEGF. If the angiogenic tissue response can be inhibited, it will be interesting to observe the degree (if any) of epidermal hyperplasia in such a setting. Similarly by injecting engrafted psoriatic plaques with inhibitory antibodies against avb3 integrin or VEGF it could be determined whether the angiogenesis could be reversed; and if it could, whether it would have any effect on the extent of keratinocyte hyperplasia in the overlying epidermis. The reagents available that antagonize or disrupt angiogenic blood vessels do so by inducing apoptosis of the endothelial cells (Brooks et al, 1994). An alternative approach to using monoclonal antibodies would be to use cytokines such as IL12 and IL-18 to attack the hyperplastic endothelial cells, and then observe the net effect on the psoriatic tissue (Coughlin et al, 1998). Thus, by taking this approach, investigators will ®nally be able to determine if the angiogenic response in the dermis is of primary or secondary importance to the genesis and propagation of psoriatic plaques. Recent discoveries related to the positive and negative regulation of blood vessel growth have propelled the ®eld of angiogenesis to the forefront of biomedical research. While it is clear that the majority of investigators are seeking to apply this new knowledge to cancer therapy and to understanding tumorigenesis, it is likely that these advances will impact our perspectives towards other disease processes such as psoriasis, rheumatoid arthritis, and many more medical conditions (Folkman, 1995). Even though a number of exciting naturally occurring and synthetic negative regulators have been discovered such as angiostatin, it will be important to identify their target receptor(s) that are responsible for mediating the antiangiogenic tissue response. While many cellular and molecular mediators of angiogenesis in psoriasis have been elucidated, many questions remain as discussed in the previous section. Nonetheless, with the development of the SCID mouse animal model as a validated and useful experimental tool, as well as the rapid advances made by investigators studying other disease processes (Eliceiri and Cheresh, 1999), it is likely that progress in understanding and treating psoriasis by attacking the vasculature will be part of our therapeutic strategy in the not so distant future. REFERENCES Bacharach-Buchies M, Panz B, Elgammal S, Aver T, Altmeyer P: The elongation of psoriatic capillaries, the result of epidermal hyperplasia, not of angiogenesis. J Invest Dermatol 103:263, 1994 Banchereau J, Steinman RM: Dendritic cells and the control of immunocytes. Nature 392:245±251, 1998 Barker JNWN, Mitra RS, Grif®ths CEM, Dixit V, Nickoloff BJ: Keratinocytes as initiators of in¯ammation: a unifying explanation for diverse array of environmental stimuli to produce cutaneous in¯ammation. Lancet 337:211±214, 1991 Boehm U, Klamp T, Grout M, Howard JC: Cellular response to interferon-gamma. Ann Rev Immunol 15:791±795, 1997 Boehncke WH, Dressel D, Zollner TM, Kaufmann R: Pulling the trigger on psoriasis. Nature 379:777, 1996 Bonish B, Jullien D, Outronc Y, et al: Overexpression of CDld by keratinocytes in psoriasis and CDld-dependent IFN-g production by NK-T cells. J Immunol 165:4076±4085, 2000 Braverman IM, Sibley J: Role of the microcirculation in the treatment and pathogenesis of psoriasis. J Invest Dermatol 78:12±17, 1982 Braverman IM, Yen A: Ultrastructure of the capillary loops in the dermal papillae of psoriasis. J Invest Dermatol 68:53±60, 1977 Brooks PC, Clark RAF, Cheresh DA: Requirement of vascular integrin avb3 for angiogenesis. Science 264:569±571, 1994 Brooks PC, Montgomery AMP, Rosenfeld M, Reisfeld RA, Hu T, Klier G, Cheresh DA: Integrin avb3 antagonists promote tumor regression by inducing apoptosis of angiogenic blood vessels. Cell 79:1157±1164, 1994 Christo®dou-Solonidou M, Bridges M, Murphy GF, Albelda SM, DeLisser HM: Expression and function of endothelial cell av integrin receptors in woundinduced human angiogenesis in human skin/SCID mouse chimeras. Am J Pathol 151:975±983, 1997

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