Effect of endothelin-1 on bovine luteal cell function: role in prostaglandin F2alpha-induced antisteroidogenic action

Endothelin-1 (ET-1) a vasoactive peptide, is synthesized and secreted by endothelial cells. In the bovine corpus luteum (CL), endothelial cells constitute a major proportion (53.5%) of total CL cells. This study was designed to examine the effects of ET-1 on bovine luteal cell functions and its involvement in the action of PGF2alpha. To better define the cells implicated in this process, we used CL slices, whole CL-derived cells, and steroidogenic large (LLC) and small (SLC) luteal-like cells. High affinity binding sites for ET-1 (K(d), approximately 0.3 x 10(-9)) were present in both steroidogenic luteal cells. The binding affinity of ET-1 was 3 orders of magnitude higher than that of ET-3, and a selective ETA receptor antagonist (BQ123) competed similarly to ET-1, suggesting the presence of ETA receptors. The lack of effect of ET-3 on CL-derived cells further supported this conclusion. Both basal progesterone secretion and bovine LH (5 ng/ml)-stimulated progesterone secretion from CL-derived cells were significantly inhibited by ET-1 in a dose-dependent manner, whereas preincubation of these cells with ETA receptor antagonist prevented the inhibitory effect of added ET-1. Incubation of LLC with 10(-8) M ET-1 inhibited their progesterone secretion (114.8 vs. 176.7 ng/10(5) cells-20 h; P < 0.05). On the other hand, ET-1 did not affect progesterone production from SLC despite the presence of ET-binding sites. PGF2alpha only inhibited LH-stimulated progesterone secretion by luteal slices. This antisteroidogenic effect of PGF2alpha could be prevented by the addition of a selective ETA receptor antagonist. Luteal tissue and microvascular endothelial cells isolated from bovine CL produced ET-1; in contrast, the peptide was undetectable in the culture medium or in cell extracts of either LLC or SLC. These data support the concept that ET-1 may play a paracrine regulatory role in bovine luteal function and propose a novel role for this peptide in mediating PGF2alpha-induced luteal regression.     

Macrophages, cell proliferation, and cell death in the human menstrual corpus luteum

We studied the presence and numbers of macrophages in the different compartments of the human menstrual corpus luteum (CL) in relation to the proliferative activity and apoptosis in luteal cells. Macrophages were recognized by immunohistochemical demonstration of the lysosome-associated glycoprotein CD68, and proliferating cells by immunohistochemical detection of the cell cycle-related protein Ki67 and by counting mitotic cells. In general, changes in the number of macrophages were parallel to the functional activity of the CL. Macrophage numbers increased up to the end of the early luteal phase, remained relatively unchanged during the midluteal phase, and decreased at the late luteal phase. Furthermore, macrophages showed prominent morphological changes during the cycle. They showed round or elongated cytoplasm during the early and late luteal phases, and dendritic features in the midluteal phase. Proliferating cells were very abundant on Days 15-16 and showed a significant decrease thereafter. Most proliferating cells corresponded to stromal (mainly vascular) cells. However, about 5% of granulosa-lutein cells and about 15% of theca-lutein cells were proliferating during the early and midluteal phases. Regression of the CL at the late luteal phase was associated with both a decrease in the number of proliferating cells and an increase in the number of apoptotic cells, which were highly increased on Days 25-27 of the cycle. The number of macrophages was not related to cell proliferation nor to cell death during the luteal phase. The observed changes in both macrophage number and morphology suggest the existence of a bidirectional communication between macrophages and steroidogenic cells in the human CL, or regulation of both cell populations by similar mechanisms.     

Differential gene expression within the ovine corpus luteum: identification of secreted protein acidic and rich in cysteine as a major secretory product of small but not large luteal cells

Limited information is available regarding secretory proteins of the corpus luteum (CL), and the potential local role these proteins may play in control of luteal function. An ovine small luteal cell complementary DNA library was immunologically screened with a polyclonal antiserum generated against small cell secretory proteins. A relatively abundant complementary DNA (approximately 2.1 kb) encoding a calcium binding glycoprotein Secreted Protein Acidic and Rich in Cysteine (SPARC) was isolated. Production of SPARC protein by ovine luteal cells was confirmed by immunoprecipitating it from labeled culture medium. Expression of SPARC messenger RNA (mRNA) was examined within CL collected on days 3, 7, 10, 13, and 16 post estrus (n = 4, 4, 4, 3, and 4 respectively), and within pools of purified small (n = 3) and large (n = 4) luteal cells by Northern and dot blot analysis. Amounts of SPARC mRNA increased during the early luteal phase, peaked by day 7 (P < 0.05) and subsequently declined on days 10 and 13 (P < 0.05). SPARC mRNA content was significantly higher in the small than in the large cells (P < 0.003). In situ hybridization showed that SPARC mRNA was localized to the thecal layer of Graafian follicles and to day 3 and day 10 CL. Within CL, immunohistochemistry indicated that SPARC protein was associated with small luteal cells (spindle shaped, avg = 17 microM in diameter) but not with large cells. This specific localization to small cells was confirmed by colocalization of SPARC with 3 beta-hydroxysteroid dehydrogenase. We conclude that SPARC is a major secretory product of small steroidogenic luteal cells of the ovine CL. As SPARC is known to modulate many aspects of tissue reorganization, expression by small luteal cells may play a role in regulation of CL maturation.     

Nutritional support for the patient with pancreatobiliary disease

The mechanisms which regulate adrenocortical steroidogenesis in differentiated parenchymal cells have been studied in great detail. However, the stem cells that are responsible for regeneration of the adult cortex have never been identified or isolated, and their characteristics are unknown. We have developed a tissue culture system that supports the simultaneous proliferation and differentiation of steroidogenic adrenocortical cells. Utilizing density gradient separation, a cell population composed of a mixture of stromal, endothelial and parenchymal cells (MIX) was isolated from the adult rat adrenal cortex. In primary culture, MIX populations formed high saturation density multilayers from which rounded cells emerged. These cells proliferated, contained lipid, and expressed the steroidogenic enzymes delta 5,3 beta-hydroxysteroid dehydrogenase and cholesterol side-chain cleavage cytochrome P-450scc. After selective passaging, rounded MIX-derived cells retained their steroidogenic potential, even in the absence of trophic hormone treatment. In contrast, parenchymal cells obtained from the zonae fasciculata (FASC) and glomerulosa (GLOM) respectively, formed homogeneous monolayers in primary culture, gradually de-differentiated, and no longer responded to trophic hormone treatment after passaging. Thus, primary MIX cultures provided a microenvironment that resulted in the production of adrenocortical cells with stem cell-like qualities. These cultures provide for the first time, a system for the identification of specific inducers that are responsible for both adrenocortical cytogenesis and its associated proliferation and steroidogenic differentiation.     

Postpubertal growth and development of the face in unilateral cleft lip and palate as compared to the pubertal period: a longitudinal study

In adult tissues, capillary growth (angiogenesis) occurs normally during tissue repair, such as in the healing of wounds and fractures. Inappropriate capillary growth is associated with various pathological conditions, including tumour growth, retinopathies, haemangiomas, fibroses and rheumatoid arthritis in the case of rampant capillary growth, and nonhealing wounds and fractures in the case of inadequate capillary growth. The female reproductive organs exhibit marked, periodic growth and regression, accompanied by equally striking changes in their rates of blood flow. It is not surprising, therefore, that they are some of the few adult tissues in which angiogenesis occurs as a normal process. Ovarian follicles and corpora lutea have been shown to contain and produce angiogenic factors. These angiogenic factors appear to be heparin-binding and to belong to the fibroblast growth factor (FGF) and vascular endothelial growth factor (VEGF) families of proteins. In addition, factors regulating gap junctional communication may play a critical role in coordinating the interactions between luteal vascular and nonvascular tissues. Further elucidation of the specific physiological roles of these factors in follicular and luteal growth, development and function will ultimately lead to improved methods for regulating fertility in mammals.     

Endothelial cell proliferation follows the mid-cycle luteinizing hormone surge, but not human chorionic gonadotrophin rescue, in the human corpus luteum

The corpus luteum is essential for the maintenance of early pregnancy in women. Angiogenesis may be one factor involved in luteal rescue. The aim of this study was to determine the changes in endothelial cell proliferation throughout the luteal phase and in human chorionic gonadotrophin (HCG)-simulated early pregnancy. Human corpora lutea obtained throughout the luteal phase and in simulated early pregnancy were immunostained with antibodies for endothelial and proliferating cells. Number and distribution of endothelial and proliferating cells were examined. Endothelial cells were least abundant in the early luteal phase, increasing in the mid-luteal phase (P < 0.03). Endothelial numbers did not differ significantly between the late and the rescued corpora lutea. Endothelial cell proliferation was greatest in the early luteal phase and continued at a lower level during later stages. Simulated early pregnancy resulted in no change in endothelial cell proliferation. These results showed that a high degree of endothelial cell proliferation is associated with formation of the human corpus luteum. Unchanging levels of proliferation following HCG treatment (for 5-8 days from day 12 to day 16 post-ovulation, at 125 IU to 16,000 IU, following a daily doubling of dose) suggest that alternative processes are involved during luteal rescue.     

Macrophages are the major source of tumor necrosis factor alpha in the porcine corpus luteum

This study was designed to determine the source of tumor necrosis factor (TNF) alpha within the porcine corpus luteum (CL). 1) Sections of frozen or paraffin-embedded CL from various stages of the estrous cycle were incubated with the following primary antibodies: anti-human recombinant TNFalpha, anti-porcine macrophage-specific antigen, or anti-alpha-actin (marker of pericyte and smooth muscle cells). Dolichos biflorus lectin-peroxidase was used as an endothelial cell label. Positive immunostaining for TNFalpha was apparent in porcine CL throughout the estrous cycle. TNFalpha immunoreactivity was primarily localized in cells along septal/vascular tracts, and exhibited spatial and temporal distribution similar to that of cells labeled with anti-macrophage antibodies. Large luteal cells exhibited weak staining for TNFalpha in paraffin sections, whereas microvascular endothelial cells were consistently negative in both frozen and paraffin sections. 2) Enriched subpopulations of macrophages, endothelial cells, and large and small luteal cells were isolated by density gradient and immunomagnetic bead separation techniques. TNFalpha secretion by each subpopulation was determined by measuring bioactive TNFalpha in incubation media using a specific in vitro bioassay. Macrophage subpopulations secreted up to 100-fold greater quantities of bioactive TNFalpha (up to 400 pg/10(6) cells) than did other subpopulations. In contrast, endothelial cell and small luteal cell subpopulations released very small amounts (< 8 pg/10(6) cells) of bioactive TNFalpha. Large luteal cells secreted slightly greater amounts of TNFalpha (10-15 pg/10(6) cells). Local macrophages appear to be the primary source of TNFalpha in the porcine CL.     

The effect of luteal "rescue" on the expression and localization of matrix metalloproteinases and their tissue inhibitors in the human corpus luteum

Luteolysis is associated with tissue remodeling probably involving the matrix metalloproteinases (MMPs) and their specific tissue inhibitors (TIMPs). This study investigated the expression and localization of the major MMPs and TIMPs in the human corpus luteum throughout the luteal phase and after luteal rescue with hCG. Corpora lutea (n = 9) were collected at hysterectomy and were dated by serial urinary LH estimation. In addition, corpora lutea (n = 3) were collected from women who had received daily doubling doses of hCG to mimic the hormonal changes of early pregnancy. MMP-1, MMP-2, MMP-9, TIMP-1, TIMP-2, and TIMP-3 were investigated by zymography, reverse zymography, Northern blotting, and in situ hybridization. There was no change in the expression of MMP-1, TIMP-1, and TIMP-2 throughout the luteal phase or after luteal rescue. Little TIMP-3 could be detected in the corpus luteum. MMP-9 activity peaked in the early and late luteal phase. The expression and activity of MMP-2 were maximal in the late luteal phase. Exposure to hCG during luteal rescue in vivo was associated with a reduction (P < 0.05) in the expression and activity of MMP-2. Messenger ribonucleic acids (mRNAs) for MMP-1, MMP-2, and TIMP-2 were localized to the connective tissue stroma and the thecal-lutein cells of the corpus luteum. In contrast, TIMP-1 mRNA was localized to the granulosa-lutein cells, and MMP-9 mRNA was expressed in scattered cells within the steroidogenic and nonsteroidogenic cell layers. In conclusion, during maternal recognition of pregnancy, hCG prevents the normal increase in MMP-2 in the late luteal phase. MMPs can function in an environment containing large amounts of TIMP-1, as they have a different cellular localization.     

Expression of the angiogenic factors, basic fibroblast growth factor and vascular endothelial growth factor, in the ovary

In adult tissues, vascular growth (angiogenesis) occurs normally during tissue repair, such as in the healing of wounds and fractures. Inappropriate vascular growth is associated with various pathological conditions. These conditions include tumor growth, retinopathies, hemangiomas, fibroses, and rheumatoid arthritis in the case of rampant vascular growth and nonhealing wounds and fractures in the case of inadequate vascular growth. The female reproductive organs exhibit dramatic, periodic growth and regression, accompanied by equally dramatic changes in their rates of blood flow. Thus, it is not surprising that they are some of the few adult tissues in which angiogenesis occurs as a normal process. Ovarian follicles and corpora lutea contain and produce angiogenic factors. These angiogenic factors bind heparin and seem to belong to the fibroblast growth factor (FGF) and vascular endothelial growth factor (VEGF) families of proteins. Based on our studies of the pattern of expression of FGF and its major receptors in bovine, ovine, and porcine corpora lutea, we have suggested that FGF may influence not only luteal cell proliferation but also cell death, thereby regulating cell turnover in the luteal vascular and nonvascular compartments. In addition, we recently have shown that luteal expression of VEGF is greatest during the early luteal phase, coincident with luteal vascularization. Moreover, VEGF is present exclusively in luteal connective tissue and perivascular (arteriolar smooth muscle and capillary pericyte) cells. In fact, the first thecal-derived cells to invade the granulosa-derived regions immediately after ovulation seem to be VEGF-containing pericytes. We have therefore hypothesized that ovarian pericytes play a key role in vascularization of developing follicles and corpora lutea. Further understanding of the specific physiological roles of these factors in follicular and luteal growth, development, and function will ultimately lead to improved methods of regulating fertility.     

Prominent expression of glial cell line-derived neurotrophic factor in human skeletal muscle

Natural killer (NK) cells are well recognized as cytolytic effector cells of the innate immune system. In the past several years, the structure and function of NK cell receptors for the major histocompatibility complex (MHC) class I molecules and other ligands have been the subject of extensive studies. These studies. These studies have focused largely on the mechanisms of target cell recognition for lysis. Another aspect of NK cell function that seems to be underappreciated is their role in immune regulation. Since NK cells produce a number of immunologically relevant cytokines, it has been suggested that these cells may modulate the development of the adaptive immune response. But, is it the only mechanism by which NK cells interact with cells involved in the induction of antigen-specific responses? This article reviews some older and more recent studies and attempts to place NK cells in the context of potent immune regulators of T cell responses.     

Identification and distribution of tumor necrosis factor alpha receptors in pig corpora lutea

This study examined whether the pig CL contains specific tumor necrosis factor alpha (TNF alpha) receptors and compared the binding affinities and capacities of small and large cell membranes. Aliquots of membranes, isolated from intact or dispersed luteal tissue, were homogenized, and membrane protein content was quantified. Luteal membranes were assayed for specific TNF alpha binding by displacement analysis, with use of [125I]TNF alpha and varying concentrations of unlabeled TNF alpha. Preliminary experiments demonstrated that TNF alpha binding was maximal after incubation at 22 degrees C for 180 min. In addition, [125I]TNF alpha binding was displaced by TNF alpha, but not by other cytokines. Small cell membranes contained a TNF alpha binding site with an affinity (Kd = 11.6-19 nM) different (p < 0.05) from that of the binding site on large cell membranes (Kd = 56.2-99.6 nM). TNF alpha binding capacities were similar in small and large cell membranes. These data demonstrate that pig CL contain specific, saturable TNF alpha binding sites. The higher-affinity binding sites were localized in the small cell population, which contains predominantly endothelial cells and small luteal cells, suggesting that TNF alpha acts primarily on one or both of these cell types within the CL.     

Structure and functions of the endothelium

The vascular endothelium is a dynamic interface between blood and tissues, which releases vasoconstrictors or vasodilators regulating the vascular tone. The endothelium modulates the balance between thrombosis and haemorrhage. Activated endothelium may produce tissue factor which triggers the coagulation cascade. In different tissues, the endothelial cells become specialised and may participate to the immune response and inflammation. Various metabolic or immune stimuli may alter endothelial cell functions, induce leukocyte adhesion through expression of specialised molecules and modify the release of fibrinolytic agents, cytokines, and growth factors.     

Gap junctional proteins, connexin 26, 32, and 43 in sheep ovaries throughout the estrous cycle

Ovarian follicles from days 13, 14, 15, and 16 and corpora lutea (CL) from days 2, 4, 8, 12, and 15 of the estrous cycle were evaluated for the presence of connexins by immunohistochemistry. In addition, CL from days 5, 10, and 15 of the estrous cycle were used for immunofluorescent detection of Cx43 followed by image analysis, and for Western immunoblot. In all tissues, staining for all connexins appeared punctate, indicating the presence of assembled gap junctions. Cx26 was present in the ovarian surface epithelium, stroma, and blood vessels within the stroma and hilus, and in the CL. In healthy antral follicles, Cx26 was present only in the theca layer, whereas Cx43 was present in granulosa and theca layers. In the majority of atretic follicles, connexins were not detected, but in 13% of the atretic follicles, Cx43 was present in the theca layer. Cx32 was detected in the blood vessels of ovarian stroma and in the CL, and Cx43 was detected in the CL. Localization and/or expression of connexins depended on stage of luteal development. Western analysis demonstrated that expression of Cx32 in luteal tissues was similar across the estrous cycle. The area of positive staining for Cx43 and expression of Cx43 in luteal tissues decreased (p < 0.05) as the estrous cycle progressed. The pattern of expression of connexins indicates that gap junctional proteins may be important in the regulation of folliculogenesis and follicular atresia, as well as growth, differentiation, and regression of the CL.     

Bovine ovarian non-genomic progesterone binding sites: presence in follicular and luteal cell membranes

We have shown recently that the bovine corpus luteum (CL) possesses specific luteal cell surface membrane binding sites for progesterone. We have now confirmed and extended these observations to compare the subcellular distribution of these binding sites in developing, mature and regressing CL. The median buoyant densities of luteal progesterone binding sites from early-, mid- and late-luteal phase CL were similar (though three of five density profiles for late-luteal phase CL showed association of steroid binding with a fraction with a lower density), and clearly resolved from nuclear, mitochondrial, lysosomal, peroxisomal, Golgi-endoplasmic reticulum-lysosomal and smooth endoplasmic reticulum markers. Specific binding of [3H]progesterone overlapped with the distributions of 5'-nucleotidase and luteinizing hormone receptor (luteal cell surface membrane markers) in both control and digitonin-treated gradients at all stages of the luteal phase. Since steroidogenic 'large luteal' and 'small luteal' cells of the CL are derived from the granulosa cells (GC) and theca of the preovulatory follicle, we also investigated whether similar receptors were present in the follicle, and describe for the first time specific membrane binding sites for progesterone in purified GC and thecal membranes from healthy bovine follicles of different sizes. Specific binding increased linearly with GC and thecal membrane protein concentration; however, it was detectable only when digitonin was included in the binding incubation. Binding sites were specific for progesterone; unlabelled progesterone competed for [3H]progesterone binding at low concentrations (IC50, 35 and 31 nmol/l) compared with testosterone (IC50, 905 and 870 nmol/l) and delta4-androstenedione (IC50, 1050 and 660 nmol/l) for GC and thecal receptors respectively. In contrast, oestradiol, oestrone, pregnenolone, cortisol, cholesterol, and a genomic progesterone receptor antagonist, RU486, competed poorly. Steroid binding was present in GC and thecal membranes of follicles of all sizes, but [3H]progesterone binding to GC membranes decreased significantly with increasing follicle size (P<0.02), perhaps indicating developmental regulation of GC membrane non-genomic progesterone receptors in the preovulatory bovine follicle. We suggest that these membrane steroid receptors may be involved in the autocrine/paracrine regulation of follicular function by progesterone.     

The pathogenesis of Graves' disease

Graves' disease may prove to be due to a genetic defect in immune surveillance or control, which then permits a randomly mutating thyroid-directed clone of T-lymphocytes to survive, interact with previously normal thyroid cell membrane antigen, and set up a localized cell-mediated immune response. The T-lymphocytes so interacting then cooperate with and direct groups of B-lymphocytes, which consequently produce TSI's: the stimulation of the thyroid parenchymal cells is a direct result of TSI stimulation. PHA stimulates Graves' lymphocytes to produce TSI's by first stimulating T-lymphocytes, which in turn interact with B-lymphocytes, which produce the TSI's. Normal thyroid antigen can stimulate sensitized lymphocytes to produce TSI's, which in turn can interact with normal thyroid cell membranes. This and other evidence suggests that there need not be any thyroidal antigenic change necessary to initiate hyperthyroidism. Stress may be related to the immunologic induction of the disease; it appears that remissions, other than those due to thyroid cell destruction, are immunologic. Exophthalmos may also prove to be an autoimmune disorder resulting from a closely related, but possibly not identical, defect in immunologic surveillance. Other related autoimmune diseases may result from similar related defects in immunologic surveillance, as well as from the random mutation of the appropriate forbidden clone.     

Prostaglandin f2alpha treatment in vivo, but not in vitro, stimulates protein kinase C-activated superoxide production by nonsteroidogenic cells of the rat corpus luteum

Luteal regression is associated with the generation of reactive oxygen species (ROS). To determine the nature of the ROS generator, cells isolated from luteinized rat ovaries were examined for ROS production using luminol-amplified chemiluminescence (LCL). Cells cultured for 2-48 h exhibited minimal LCL, but there was a significant (30- to 50-fold), rapid (maximum at 3-5 min), and dose-dependent increase in LCL in response to phorbol ester (phorbol 12-myristate 13-acetate; TPA; ED50 = 0.03 microM) and diacylglycerol (1,2-dioctanoyl-glycerol; ED50 = 30 microM). The TPA-induced response was cell number dependent and was virtually abolished by superoxide dismutase, freezing, or heating (95 degrees C for 5 min). Zymosan, known to induce a phagocytic response in leukocytes, stimulated a superoxide (O2-.) response with a slow onset (maximum at 40 to 60 min) and a maximum about one third of that observed for TPA. The response to TPA and zymosan was inhibited by the NADPH/NADH-oxidase inhibitor, diphenylene iodonium (ID50 = 5 microM for TPA), but not by the mitochondrial inhibitors, potassium cyanide, rotenone, or sodium azide. Fractionation of cells by centrifugal elutriation showed that TPA-stimulated O2-. production coeluted with the nonsteroidogenic cells and that little, if any, O2-. generation coeluted with the steroidogenic cells. Cells isolated 1, 2, and 4 h after in vivo treatment with a luteolytic dose of prostaglandin F2alpha (PGF2alpha) showed a significant increase in TPA-stimulated O2-. production at 2 h, whereas luteal cells or corpora lutea incubated directly with 1 microM PGF2alpha did not show any increase in response. Corpora lutea isolated from naturally regressed ovaries (18 days after ovulation) showed a significantly elevated level of TPA-stimulated O2-. production. In conclusion, there is a superoxide generator in luteinized ovaries that is activated through a protein kinase C pathway, localized in nonsteroidogenic cells, transiently increased during PGF2alpha-induced luteolysis in vivo, and elevated during natural luteal regression.     

Growth regulation of vascular cells by cytokines]

Cytokines with stimulatory or inhibitory activities for vascular cells are reviewed. Directly or via humoral factors, vascular endothelial cells interact with blood cells, such as lymphocytes, neutrophils and platelets, while smooth muscle cells do so with inflammatory cells. Various cytokines, including IL-1, 6, 7, 8, GM-, G-, M-CSF, a, b-FGF, PDGF, TGF beta, PAF, PA, PAI-1, cell adhesion molecules and endothelin are produced by endothelial cells and/or smooth muscle cells, and in turn they and cytokines produced by blood cells, act as modulators of growth or function of the vascular cells under some physico-pathological states. Vascular cells, especially, endothelial cells might thus be involved in cytokine network.     

Bulge ductility of several occlusal contact measuring paper-based and plastic-based sheets

During angiogenesis new blood vessels sprout from an existing vascular bed. This is a prerequisite for tumor growth beyond a certain size and for metastasis formation. Tumors produce a number of cytokines. The development of in vivo bio-assays for angiogenesis and in vitro analysis of endothelial cells permits characterization of these different cytokines concerning their role in angiogenesis. Some cytokines act mitogenically on endothelial cells, others have chemotactic activity or induce tube formation and some have multiple functions. A few factors are chemotactic for macrophages, infiltrating the tumor and secreting further angiogenic cytokines. Another important role in the process of angiogenesis is played by the extracellular matrix. Proteases secreted by all cell types involved (tumor cell, endothelial cell, macrophages) degrade the extracellular matrix, thereby releasing and activating angiogenic factors sequestered in the extracellular matrix. Thus tumor cells, macrophages and extracellular matrix release cytokines which together act on endothelial cells, resulting in the growth and infiltration of new blood vessels into the tumor.     

Immunity at the surface: homeostatic mechanisms of the skin immune system

The coordinated function of multiple epidermal and dermal cell populations allows the skin immune system to respond rapidly and effectively to a wide variety of insults occurring at the interface of the organism and its environment. Keratinocytes are the first line of defense in the skin immune system, and keratinocyte-derived cytokines are pivotal in mobilizing leukocytes from blood and signaling other cutaneous cells. Cytokine-mediated cellular communication also enables dermal fibroblasts and endothelial cells lining the cutaneous vasculature to participate in immune and inflammatory responses. Skin is an important site for antigen presentation, and both epidermal Langerhans cells and dermal dendritic cells play pivotal roles in T cell-mediated immune responses to antigens encountered in skin. Proinflammatory signaling pathways are necessarily balanced by a variety of regulatory pathways that help maintain the homeostatic functioning of the skin immune system.     

TGF-beta and the endothelium during immune injury

During immune injury, activation of endothelial cells by inflammatory cytokines stimulates leukocyte adhesion to the endothelium, turns the endothelium from an anticoagulant surface to one that is frankly procoagulant, and results in the release of vasoactive mediators and growth factors. Cytokine activation of endothelial cells also results in increased endothelial cell TGF-beta 1 synthesis and enhanced activation of latent TGF-beta, the latter involving a shift of plasmin production from the apical to subendothelial surface. In cytokine-stimulated endothelial cells, TGF-beta hinders leukocyte adhesion and transmigration via inhibition of IL-8 and E-selectin expression. TGF-beta also profoundly diminishes cytokine-stimulated inducible nitric oxide synthase production and instead augments endothelial nitric oxide synthase expression. Thus, some of the TGF-beta actions on endothelium during immune activation can viewed as immunosuppressive. TGF-beta also influences mechanisms of vascular remodeling during the healing phase of immune injury. It stimulates PDGF-B synthesis by endothelial cells, causes bFGF release from subendothelial matrix, and promotes VEGF synthesis by non-endothelial cells. Together these mediators control angiogenesis, a critical component of the vascular repair phenomenon. Further, endothelial cell derived PDGF-B and bFGF influence the proliferation and migration of neighboring cells. Thus, endothelial cells and TGF-beta actions on the endothelium play important roles both during the initial phase of immune injury and during the later remodeling phase.